Utkan Demirci

All Publications

• Co-axial hydrogel spinning for facile biofabrication of prostate cancer-like 3D models. Biofabrication Guimaraes, C. F., Liu, S., Wang, J., Purcell, E., Ozedirne, T., Ren, T., Aslan, M., Yin, Q., Reis, R. L., Stoyanova, T., Demirci, U. 2024

  Abstract

  Glandular cancers are amongst the most prevalent types of cancer, which can develop in many different organs, presenting challenges in their detection as well as high treatment variability and failure rates. For that purpose, anticancer drugs are commonly tested in cancer cell lines grown in 2D tissue culture on plastic dishes in vitro, or in animal models in vivo. However, 2D culture models diverge significantly from the 3D characteristics of living tissues and animal models require extensive animal use and time. Glandular cancers, such as prostate cancer - the second leading cause of male cancer death - typically exist in co-centrical architectures where a cell layer surrounds an acellular lumen. Herein, this spatial cellular position and 3D architecture, containing dual compartments with different hydrogel materials, is engineered using a simple co-axial nozzle setup, in a single step utilizing prostate as a model of glandular cancer. The resulting hydrogel soft structures support viable prostate cancer cells of different cell lines and enable over-time maturation into cancer-mimicking aggregates surrounding the acellular core. The biofabricated cancer mimicking structures are then used as a model to predict the inhibitory efficacy of the poly ADP ribose polymerase (PARP) inhibitor, Talazoparib, and the antiandrogen drug, Enzalutamide, in the growth of the cancer cell layer. Our results show that the obtained hydrogel constructs can be adapted to quickly obtain 3D cancer models which combine 3D physiological architectures with high-throughput screening to detect and optimize anti-cancer drugs in prostate and potentially other glandular cancer types.

  View details for DOI 10.1088/1758-5090/ad2535

  View details for PubMedID 38306674

• Integrated "lab-on-a-chip" microfluidic systems for isolation, enrichment, and analysis of cancer biomarkers. Lab on a chip Surappa, S., Multani, P., Parlatan, U., Sinawang, P. D., Kaifi, J., Akin, D., Demirci, U. 2023

  Abstract

  The liquid biopsy has garnered considerable attention as a complementary clinical tool for the early detection, molecular characterization and monitoring of cancer over the past decade. In contrast to traditional solid biopsy techniques, liquid biopsy offers a less invasive and safer alternative for routine cancer screening. Recent advances in microfluidic technologies have enabled handling of liquid biopsy-derived biomarkers with high sensitivity, throughput, and convenience. The integration of these multi-functional microfluidic technologies into a 'lab-on-a-chip' offers a powerful solution for processing and analyzing samples on a single platform, thereby reducing the complexity, bio-analyte loss and cross-contamination associated with multiple handling and transfer steps in more conventional benchtop workflows. This review critically addresses recent developments in integrated microfluidic technologies for cancer detection, highlighting isolation, enrichment, and analysis strategies for three important sub-types of cancer biomarkers: circulating tumor cells, circulating tumor DNA and exosomes. We first discuss the unique characteristics and advantages of the various lab-on-a-chip technologies developed to operate on each biomarker subtype. This is then followed by a discussion on the challenges and opportunities in the field of integrated systems for cancer detection. Ultimately, integrated microfluidic platforms form the core of a new class of point-of-care diagnostic tools by virtue of their ease-of-operation, portability and high sensitivity. Widespread availability of such tools could potentially result in more frequent and convenient screening for early signs of cancer at clinical labs or primary care offices.

  View details for DOI 10.1039/d2lc01076c

  View details for PubMedID 37314731

• Systematic Analysis of Tissue-Derived and Biofluid Extracellular Vesicle miRNAs Associated with Prostate Cancer. Advanced biology Larson, J., Ozen, M. O., Kohli, M., Akin, D., Demirci, U. 2023: e2200327

  Abstract

  Extracellular vesicles (EVs) are emerging as biomarker candidates for early detection of prostate cancer. Studies compare EV-microRNA (miRNA) expression in individuals with prostate cancer (PCa) with cancer-free samples for diagnostic purposes. The aim of this study is to review miRNA signatures to investigate the overlap between miRNAs enriched in PCa tissue and miRNAs enriched in EVs isolated from subjects with PCa biofluids (i.e., urine, serum, and plasma). Signatures dysregulated in EVs from PCa biofluids and tissue are potentially associated with the primary tumor site and might be more indicative of PCa at an early stage. A systematic review of EV-derived miRNAs and a reanalysis of PCa tissue miRNA sequencing data for comparison is presented. Articles in the literature are screened for validated miRNA dysregulation in PCa and compared with TCGA primary PCa tumor data using DESeq2. This resulted in 190 dysregulated miRNAs being identified. Thirty-one eligible studies are identified, indicating 39 dysregulated EV-derived miRNAs. The top ten markers identified as significantly dysregulated in the PCa tissue dataset TCGA (e.g., miR-30b-3p, miR-210-3p, miR-126-3p, and miR-196a-5p) have a significant expression change in EVs with the same directionality in one or several statistically significant results. This analysis highlights several less frequently studied miRNAs in PCa literature.

  View details for DOI 10.1002/adbi.202200327

  View details for PubMedID 37300338

• Methods to Evaluate Changes in Mitochondrial Structure and Function in Cancer. Cancers Rickard, B. P., Overchuk, M., Chappell, V. A., Kemal Ruhi, M., Sinawang, P. D., Nguyen Hoang, T. T., Akin, D., Demirci, U., Franco, W., Fenton, S. E., Santos, J. H., Rizvi, I. 2023; 15 (9)

  Abstract

  Mitochondria are regulators of key cellular processes, including energy production and redox homeostasis. Mitochondrial dysfunction is associated with various human diseases, including cancer. Importantly, both structural and functional changes can alter mitochondrial function. Morphologic and quantifiable changes in mitochondria can affect their function and contribute to disease. Structural mitochondrial changes include alterations in cristae morphology, mitochondrial DNA integrity and quantity, and dynamics, such as fission and fusion. Functional parameters related to mitochondrial biology include the production of reactive oxygen species, bioenergetic capacity, calcium retention, and membrane potential. Although these parameters can occur independently of one another, changes in mitochondrial structure and function are often interrelated. Thus, evaluating changes in both mitochondrial structure and function is crucial to understanding the molecular events involved in disease onset and progression. This review focuses on the relationship between alterations in mitochondrial structure and function and cancer, with a particular emphasis on gynecologic malignancies. Selecting methods with tractable parameters may be critical to identifying and targeting mitochondria-related therapeutic options. Methods to measure changes in mitochondrial structure and function, with the associated benefits and limitations, are summarized.

  View details for DOI 10.3390/cancers15092564

  View details for PubMedID 37174030

• Programmable Shape Morphing Metasponge ADVANCED INTELLIGENT SYSTEMS Soto, F., Tsui, A., Surappa, S., Ahmed, R., Wang, J., Kilinc, U., Akin, D., Demirci, U. 2023

  View details for DOI 10.1002/aisy.202300043

  View details for Web of Science ID 000974241500001

• A photonic resonator interferometric scattering microscope for label-free detection of nanometer-scale objects with digital precision in point-of-use environments. Biosensors & bioelectronics Liu, L., Tibbs, J., Li, N., Bacon, A., Shepherd, S., Lee, H., Chauhan, N., Demirci, U., Wang, X., Cunningham, B. T. 2023; 228: 115197

  Abstract

  Label-free detection and digital counting of nanometer-scaled objects such as nanoparticles, viruses, extracellular vesicles, and protein molecules enable a wide range of applications in cancer diagnostics, pathogen detection, and life science research. Here, we report the design, implementation, and characterization of a compact Photonic Resonator Interferometric Scattering Microscope (PRISM) designed for point-of-use environments and applications. The contrast of interferometric scattering microscopy is amplified through a photonic crystal surface, upon which scattered light from an object combines with illumination from a monochromatic source. The use of a photonic crystal substrate for interferemetric scattering microscopy results in reduced requirements for high-intensity lasers or oil-immersion objectives, thus opening a pathway toward instruments that are more suitable for environments outside the optics laboratory. The instrument incorporates two innovative elements that facilitate operation on a desktop in ordinary laboratory environments by users that do not have optics expertise. First, because scattering microscopes are extremely sensitive to vibration, we incorporated an inexpensive but effective solution of suspending the instrument's main components from a rigid metal framework using elastic bands, resulting in an average of 28.7 dBV reduction in vibration amplitude compared to an office desk. Second, an automated focusing module based on the principle of total internal reflection maintains the stability of image contrast over time and spatial position. In this work, we characterize the system's performance by measuring the contrast from gold nanoparticles with diameters in the 10-40 nm range and by observing various biological analytes, including HIV virus, SARS-CoV-2 virus, exosome, and ferritin protein.

  View details for DOI 10.1016/j.bios.2023.115197

  View details for PubMedID 36905862

• Review of HIV Self Testing Technologies and Promising Approaches for the Next Generation. Biosensors Bacon, A., Wang, W., Lee, H., Umrao, S., Sinawang, P. D., Akin, D., Khemtonglang, K., Tan, A., Hirshfield, S., Demirci, U., Wang, X., Cunningham, B. T. 2023; 13 (2)

  Abstract

  The ability to self-test for HIV is vital to preventing transmission, particularly when used in concert with HIV biomedical prevention modalities, such as pre-exposure prophylaxis (PrEP). In this paper, we review recent developments in HIV self-testing and self-sampling methods, and the potential future impact of novel materials and methods that emerged through efforts to develop more effective point-of-care (POC) SARS-CoV-2 diagnostics. We address the gaps in existing HIV self-testing technologies, where improvements in test sensitivity, sample-to-answer time, simplicity, and cost are needed to enhance diagnostic accuracy and widespread accessibility. We discuss potential paths toward the next generation of HIV self-testing through sample collection materials, biosensing assay techniques, and miniaturized instrumentation. We discuss the implications for other applications, such as self-monitoring of HIV viral load and other infectious diseases.

  View details for DOI 10.3390/bios13020298

  View details for PubMedID 36832064

• Color-selective labyrinth-like quantum dot nanobeads enable point-of-care dual assay of of Mycotoxins SENSORS AND ACTUATORS B-CHEMICAL Hu, X., Huang, L., Wang, S., Ahmed, R., Li, P., Demirci, U., Zhang, Z. 2023; 376

  View details for DOI 10.1016/j.snb.2022.132956

  View details for Web of Science ID 000904331400004

• Label-Free Identification of Exosomes using Raman Spectroscopy and Machine Learning. Small (Weinheim an der Bergstrasse, Germany) Parlatan, U., Ozen, M. O., Kecoglu, I., Koyuncu, B., Torun, H., Khalafkhany, D., Loc, I., Ogut, M. G., Inci, F., Akin, D., Solaroglu, I., Ozoren, N., Unlu, M. B., Demirci, U. 2023: e2205519

  Abstract

  Exosomes, nano-sized extracellular vesicles (EVs) secreted from cells, carry various cargo molecules reflecting their cells of origin. As EV content, structure, and size are highly heterogeneous, their classification via cargo molecules by determining their origin is challenging. Here, a method is presented combining surface-enhanced Raman spectroscopy (SERS) with machine learning algorithms to employ the classification of EVs derived from five different cell lines to reveal their cellular origins. Using an artificial neural network algorithm, it is shown that the label-free Raman spectroscopy method's prediction ratio correlates with the ratio of HT-1080 exosomes in the mixture. This machine learning-assisted SERS method enables a new direction through label-free investigation of EV preparations by differentiating cancer cell-derived exosomes from those of healthy. This approach will potentially open up new avenues of research for early detection and monitoring of various diseases, including cancer.

  View details for DOI 10.1002/smll.202205519

  View details for PubMedID 36642804

• A Target Recycling Amplification Process for the Digital Detection of Exosomal MicroRNAs through Photonic Resonator Absorption Microscopy. Angewandte Chemie (International ed. in English) Wang, X., Shepherd, S., Li, N., Che, C., Song, T., Xiong, Y., Palm, I. R., Zhao, B., Kohli, M., Demirci, U., Lu, Y., Cunningham, B. 2023

  Abstract

  Exosomal microRNAs (miRNAs) have considerable potential as pivotal biomarkers to monitor cancer development, dis-ease progression, treatment effects and prognosis. Here, we report an efficient target recycling amplification process (TRAP) for the digital detection of miRNAs using photonic resonator absorption microscopy. We achieve multiplex digital detection with sub-attomolar sensitivity in 20 minutes, robust selectivity for single nucleotide variants, and a broad dynamic range from 1 aM to 1 pM. Compared with traditional qRT-PCR, TRAP showed similar accuracy in profiling exosomal miRNAs derived from cancer cells, but also exhibited at least 31-fold and 61-fold enhancement in the limits of miRNA-375 and miRNA-21 detection, respectively. The TRAP approach is ideal for exosomal or circulating miRNA biomarker quantification, where the miRNAs are present in low concentrations or sample volume, with potentials for frequent, low-cost, and minimally invasive point-of-care testing.

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  View details for DOI 10.1002/anie.202217932

  View details for PubMedID 36622783

• A Photonic Resonator Interferometric Scattering Microscope for Label-free Detection of Nanometer-Scale Objects with Digital Precision in Point-of-Use Environments. bioRxiv : the preprint server for biology Liu, L., Tibbs, J., Li, N., Bacon, A., Shepherd, S., Lee, H., Chauhan, N., Demirci, U., Wang, X., Cunningham, B. T. 2022

  Abstract

  Label-free detection and digital counting of nanometer-scaled objects such as nanoparticles, viruses, extracellular vesicles, and protein molecules enable a wide range of applications in cancer diagnostics, pathogen detection, and life science research. The contrast of interferometric scattering microscopy is amplified through a photonic crystal surface, upon which scattered light from an object combines with illumination from a monochromatic plane wave source. The use of a photonic crystal substrate for interference scattering microscopy results in reduced requirements for high-intensity lasers or oil-immersion objectives, thus opening a pathway toward instruments that are more suitable for environments outside the optics laboratory. Here, we report the design, implementation, and characterization of a compact Photonic Resonator Interferometric Scattering Microscope (PRISM) designed for point-of-use environments and applications. The instrument incorporates two innovative elements that facilitate operation on a desktop in ordinary laboratory environments by users that do not have optics expertise. First, because scattering microscopes are extremely sensitive to vibration, we incorporated an inexpensive but effective solution of suspending the instrument's main components from a rigid metal framework using elastic bands, resulting in an average of 28.7 dBV reduction in vibration amplitude compared to an office desk. Second, an automated focusing module based on the principle of total internal reflection maintains the stability of image contrast over time and spatial position, facilitating automated data collection. In this work, we characterize the system's performance by measuring the contrast from gold nanoparticles with diameters in the 10-40 nm range and by observing various biological analytes, including HIV virus, SARS-CoV-2 virus, exosomes, and ferritin protein.

  View details for DOI 10.1101/2022.12.13.520266

  View details for PubMedID 36561182

  View details for PubMedCentralID PMC9774210

• Acoustic fabrication of living toroids and cardiomyocyte-based hybrid biorobots Demirci, U. MARY ANN LIEBERT, INC. 2022: 149

  View details for Web of Science ID 000899413100140

• Large-Scale Functionalized Metasurface-Based SARS-CoV-2 Detection and Quantification. ACS nano Ahmed, R., Guimaraes, C. F., Wang, J., Soto, F., Karim, A. H., Zhang, Z., Reis, R. L., Akin, D., Paulmurugan, R., Demirci, U. 2022

  Abstract

  Plasmonic metasurfaces consist of metal-dielectric interfaces that are excitable at background and leakage resonant modes. The sharp and plasmonic excitation profile of metal-free electrons on metasurfaces at the nanoscale can be used for practical applications in diverse fields, including optoelectronics, energy harvesting, and biosensing. Currently, Fano resonant metasurface fabrication processes for biosensor applications are costly, need clean room access, and involve limited small-scale surface areas that are not easy for accurate sample placement. Here, we leverage the large-scale active area with uniform surface patterns present on optical disc-based metasurfaces as a cost-effective method to excite asymmetric plasmonic modes, enabling tunable optical Fano resonance interfacing with a microfluidic channel for multiple target detection in the visible wavelength range. We engineered plasmonic metasurfaces for biosensing through efficient layer-by-layer surface functionalization toward real-time measurement of target binding at the molecular scale. Further, we demonstrated the quantitative detection of antibodies, proteins, and the whole viral particles of SARS-CoV-2 with a high sensitivity and specificity, even distinguishing it from similar RNA viruses such as influenza and MERS. This cost-effective plasmonic metasurface platform offers a small-scale light-manipulation system, presenting considerable potential for fast, real-time detection of SARS-CoV-2 and pathogens in resource-limited settings.

  View details for DOI 10.1021/acsnano.2c02500

  View details for PubMedID 36125414

• Photochemical Targeting of Mitochondria to Overcome Chemoresistance in Ovarian Cancer. Photochemistry and photobiology Rickard, B. P., Overchuk, M., Obaid, G., Ruhi, M. K., Demirci, U., Fenton, S. E., Santos, J. H., Kessel, D., Rizvi, I. 2022

  Abstract

  Ovarian cancer is the most lethal gynecologic malignancy with a stubborn mortality rate of ~65%. The persistent failure of multi-line chemotherapy, and significant tumor heterogeneity, have made it challenging to improve outcomes. A target of increasing interest is the mitochondrion because of its essential role in critical cellular functions, and the significance of metabolic adaptation in chemoresistance. This review describes mitochondrial processes, including metabolic reprogramming, mitochondrial transfer, and the state of mitochondrial networks, in ovarian cancer progression and chemoresistance. The effect of malignant ascites, or excess peritoneal fluid, on mitochondrial function is discussed. The role of photodynamic therapy (PDT) in overcoming mitochondria-mediated resistance, is presented. PDT, a photochemistry-based modality, involves the light-based activation of a photosensitizer leading to the production of short-lived reactive molecular species and spatio-temporally confined photodamage to nearby organelles and biological targets. The consequential effects range from sub-cytotoxic priming of target cells for increased sensitivity to subsequent treatments, such as chemotherapy, to direct cell killing. This review discusses how PDT-based approaches can address key limitations of current treatments. Specifically, an overview of the mechanisms by which PDT alters mitochondrial function, and a summary of preclinical advancements and clinical PDT experience in ovarian cancer are provided.

  View details for DOI 10.1111/php.13723

  View details for PubMedID 36117466

• Automated Recognition of Plasmodium falciparum Parasites from Portable Blood Levitation Imaging. Advanced science (Weinheim, Baden-Wurttemberg, Germany) Deshmukh, S. S., Byaruhanga, O., Tumwebaze, P., Akin, D., Greenhouse, B., Egan, E. S., Demirci, U. 2022: e2105396

  Abstract

  In many malaria-endemic regions, current detection tools are inadequate in diagnostic accuracy and accessibility. To meet the need for direct, phenotypic, and automated malaria parasite detection in field settings, a portable platform to process, image, and analyze whole blood to detect Plasmodium falciparum parasites, is developed. The liberated parasites from lysed red blood cells suspended in a magnetic field are accurately detected using this cellphone-interfaced, battery-operated imaging platform. A validation study is conducted at Ugandan clinics, processing 45 malaria-negative and 36 malaria-positive clinical samples without external infrastructure. Texture and morphology features are extracted from the sample images, and a random forest classifier is trained to assess infection status, achieving 100% sensitivity and 91% specificity against gold-standard measurements (microscopy and polymerase chain reaction), and limit of detection of 31 parasites per µL. This rapid and user-friendly platform enables portable parasite detection and can support malaria diagnostics, surveillance, and research in resource-constrained environments.

  View details for DOI 10.1002/advs.202105396

  View details for PubMedID 35957519

• Engineered living bioassemblies for biomedical and functional material applications. Current opinion in biotechnology Guimaraes, C. F., Soto, F., Wang, J., Akin, D., Reis, R. L., Demirci, U. 2022; 77: 102756

  Abstract

  Recent breakthroughs in biofabrication of bioasemblies, consisting of the engineered structures composed of biological or biosynthetic components into a single construct, have found a wide range of practical applications in medicine and engineering. This review presents an overview of how the bottom-up assembly of living entities could drive advances in medicine, by developing tunable biological models and more precise methods for quantifying biological events. Moreover, we delve into advances beyond biomedical applications, where bioassemblies can be manipulated as functional robots and construction materials. Finally, we address the potential challenges and opportunities in the field of engineering living bioassemblies, toward building new design principles for the next generation of bioengineering applications.

  View details for DOI 10.1016/j.copbio.2022.102756

  View details for PubMedID 35930844

• Mutant KRAS regulates transposable element RNA and innate immunity via KRAB zinc-finger genes. Cell reports Reggiardo, R. E., Maroli, S. V., Halasz, H., Ozen, M., Hrabeta-Robinson, E., Behera, A., Peddu, V., Carrillo, D., LaMontagne, E., Whitehead, L., Kim, E., Malik, S., Fernandes, J., Marinov, G., Collisson, E., Brooks, A., Demirci, U., Kim, D. H. 2022; 40 (3): 111104

  Abstract

  RAS genes are the most frequently mutated oncogenes in cancer, yet the effects of oncogenic RAS signaling on the noncoding transcriptome remain unclear. We analyzed the transcriptomes of human airway and bronchial epithelial cells transformed with mutant KRAS to define the landscape of KRAS-regulated noncoding RNAs. We find that oncogenic KRAS signaling upregulates noncoding transcripts throughout the genome, many of which arise from transposable elements (TEs). These TE RNAs exhibit differential expression, are preferentially released in extracellular vesicles, and are regulated by KRAB zinc-finger (KZNF) genes, which are broadly downregulated in mutant KRAS cells and lung adenocarcinomas invivo. Moreover, mutant KRAS induces an intrinsic IFN-stimulated gene (ISG) signature that is often seen across many different cancers. Our results indicate that mutant KRAS remodels the repetitive noncoding transcriptome, demonstrating the broad scope of intracellular and extracellular RNAs regulated by this oncogenic signaling pathway.

  View details for DOI 10.1016/j.celrep.2022.111104

  View details for PubMedID 35858545

• Volbots: Volvox Microalgae-Based Robots for Multimode Precision Imaging and Therapy ADVANCED FUNCTIONAL MATERIALS Wang, J., Soto, F., Liu, S., Yin, Q., Purcell, E., Zeng, Y., Hsu, E., Akin, D., Sinclair, B., Stoyanova, T., Demirci, U. 2022

  View details for DOI 10.1002/adfm.202201800

  View details for Web of Science ID 000812853500001

• Endogenous Retroviral Elements Generate Pathologic Neutrophils in Pulmonary Arterial Hypertension. American journal of respiratory and critical care medicine Taylor, S., Isobe, S., Cao, A., Contrepois, K., Benayoun, B. A., Jiang, L., Wang, L., Melemenidis, S., Ozen, M. O., Otsuki, S., Shinohara, T., Sweatt, A. J., Kaplan, J., Moonen, J., Marciano, D. P., Gu, M., Miyagawa, K., Hayes, B., Sierra, R. G., Kupitz, C. J., Del Rosario, P. A., Hsi, A., Thompson, A. A., Ariza, M. E., Demirci, U., Zamanian, R. T., Haddad, F., Nicolls, M. R., Snyder, M. P., Rabinovitch, M. 2022

  Abstract

  RATIONALE: The role of neutrophils and their extracellular vesicles (EVs) in the pathogenesis of pulmonary arterial hypertension is unclear.OBJECTIVES: Relate functional abnormalities in pulmonary arterial hypertension neutrophils and their EVs to mechanisms uncovered by proteomic and transcriptomic profiling.METHODS: Production of elastase, release of extracellular traps, adhesion and migration were assessed in neutrophils from pulmonary arterial hypertension patients and control subjects. Proteomic analyses were applied to explain functional perturbations, and transcriptomic data were used to find underlying mechanisms. CD66b-specific neutrophil EVs were isolated from plasma of patients with pulmonary arterial hypertension and we determined whether they produce pulmonary hypertension in mice.MEASUREMENTS AND MAIN RESULTS: Neutrophils from pulmonary arterial hypertension patients produce and release increased neutrophil elastase, associated with enhanced extracellular traps. They exhibit reduced migration and increased adhesion attributed to elevated beta1integrin and vinculin identified on proteomic analysis and previously linked to an antiviral response. This was substantiated by a transcriptomic interferon signature that we related to an increase in human endogenous retrovirus k envelope protein. Transfection of human endogenous retrovirus k envelope in a neutrophil cell line (HL-60) increases neutrophil elastase and interferon genes, whereas vinculin is increased by human endogenous retrovirus k dUTPase that is elevated in patient plasma. Neutrophil EVs from patient plasma contain increased neutrophil elastase and human endogenous retrovirus k envelope and induce pulmonary hypertension in mice, mitigated by elafin, an elastase inhibitor.CONCLUSIONS: Elevated human endogenous retroviral elements and elastase link a neutrophil innate immune response to pulmonary arterial hypertension.

  View details for DOI 10.1164/rccm.202102-0446OC

  View details for PubMedID 35696338

• Acoustic Fabrication of Living Cardiomyocyte-based Hybrid Biorobots. ACS nano Wang, J., Soto, F., Ma, P., Ahmed, R., Yang, H., Chen, S., Wang, J., Liu, C., Akin, D., Fu, K., Cao, X., Chen, P., Hsu, E. C., Soh, H. T., Stoyanova, T., Wu, J. C., Demirci, U. 2022

  Abstract

  Organized assemblies of cells have demonstrated promise as bioinspired actuators and devices; still, the fabrication of such "biorobots" has predominantly relied on passive assembly methods that reduce design capabilities. To address this, we have developed a strategy for the rapid formation of functional biorobots composed of live cardiomyocytes. We employ tunable acoustic fields to facilitate the efficient aggregation of millions of cells into high-density macroscopic architectures with directed cell orientation and enhanced cell-cell interaction. These biorobots can perform actuation functions both through naturally occurring contraction-relaxation cycles and through external control with chemical and electrical stimuli. We demonstrate that these biorobots can be used to achieve controlled actuation of a soft skeleton and pumping of microparticles. The biocompatible acoustic assembly strategy described here should prove generally useful for cellular manipulation in the context of tissue engineering, soft robotics, and other applications.

  View details for DOI 10.1021/acsnano.2c01908

  View details for PubMedID 35671037

• Robotic Pill for Biomarker and Fluid Sampling in the Gastrointestinal Tract ADVANCED INTELLIGENT SYSTEMS Soto, F., Purcell, E., Ozen, M., Sinawang, P., Wang, J., Akin, D., Demirci, U. 2022

  View details for DOI 10.1002/aisy.202200030

  View details for Web of Science ID 000778957300001

• A Label-Free Electrical Impedance Spectroscopy for Detection of Clusters of Extracellular Vesicles Based on Their Unique Dielectric Properties. Biosensors Zhang, Y., Murakami, K., Borra, V. J., Ozen, M. O., Demirci, U., Nakamura, T., Esfandiari, L. 2022; 12 (2)

  Abstract

  Extracellular vesicles (EVs) have gained considerable attention as vital circulating biomarkers since their structure and composition resemble the originating cells. The investigation of EVs' biochemical and biophysical properties is of great importance to map them to their parental cells and to better understand their functionalities. In this study, a novel frequency-dependent impedance measurement system has been developed to characterize EVs based on their unique dielectric properties. The system is composed of an insulator-based dielectrophoretic (iDEP) device to entrap and immobilize a cluster of vesicles followed by utilizing electrical impedance spectroscopy (EIS) to measure their impedance at a wide frequency spectrum, aiming to analyze both their membrane and cytosolic charge-dependent contents. The EIS was initially utilized to detect nano-size vesicles with different biochemical compositions, including liposomes synthesized with different lipid compositions, as well as EVs and lipoproteins with similar biophysical properties but dissimilar biochemical properties. Moreover, EVs derived from the same parental cells but treated with different culture conditions were characterized to investigate the correlation of impedance changes with biochemical properties and functionality in terms of pro-inflammatory responses. The system also showed the ability to discriminate between EVs derived from different cellular origins as well as among size-sorted EVs harbored from the same cellular origin. This proof-of-concept approach is the first step towards utilizing EIS as a label-free, non-invasive, and rapid sensor for detection and characterization of pathogenic EVs and other nanovesicles in the future.

  View details for DOI 10.3390/bios12020104

  View details for PubMedID 35200364

• A Cell Culture Chip with Transparent, Micropillar-Decorated Bottom for Live Cell Imaging and Screening of Breast Cancer Cells. Micromachines Ermis, M., Antmen, E., Kuren, O., Demirci, U., Hasirci, V. 1800; 13 (1)

  Abstract

  In the recent years, microfabrication technologies have been widely used in cell biology, tissue engineering, and regenerative medicine studies. Today, the implementation of microfabricated devices in cancer research is frequent and advantageous because it enables the study of cancer cells in controlled microenvironments provided by the microchips. Breast cancer is one of the most common cancers in women, and the way breast cancer cells interact with their physical microenvironment is still under investigation. In this study, we developed a transparent cell culture chip (Ch-Pattern) with a micropillar-decorated bottom that makes live imaging and monitoring of the metabolic, proliferative, apoptotic, and morphological behavior of breast cancer cells possible. The reason for the use of micropatterned surfaces is because cancer cells deform and lose their shape and acto-myosin integrity on micropatterned substrates, and this allows the quantification of the changes in morphology and through that identification of the cancerous cells. In the last decade, cancer cells were studied on micropatterned substrates of varying sizes and with a variety of biomaterials. These studies were conducted using conventional cell culture plates carrying patterned films. In the present study, cell culture protocols were conducted in the clear-bottom micropatterned chip. This approach adds significantly to the current knowledge and applications by enabling low-volume and high-throughput processing of the cell behavior, especially the cell-micropattern interactions. In this study, two different breast cancer cell lines, MDA-MB-231 and MCF-7, were used. MDA-MB-231 cells are invasive and metastatic, while MCF-7 cells are not metastatic. The nuclei of these two cell types deformed to distinctly different levels on the micropatterns, had different metabolic and proliferation rates, and their cell cycles were affected. The Ch-Pattern chips developed in this study proved to have significant advantages when used in the biological analysis of live cells and highly beneficial in the study of screening breast cancer cell-substrate interactions in vitro.

  View details for DOI 10.3390/mi13010093

  View details for PubMedID 35056257

• Size- and density-dependent acoustic differential bioassembly of spatially-defined heterocellular architecture. Biofabrication Gu, L., Jiang, S., Xu, X., Wang, J., Xu, F., Fan, H., Shang, J., Liu, K., Demirci, U., Chen, P. 2022; 15 (1)

  Abstract

  Emerging acoustic bioassembly represents an attractive strategy to build cellular closely-packed organotypic constructs in a tunable manner for biofabrication. However, simultaneously assemble heterogeneous cell types into heterocellular functional units with spatially-defined cell arrangements, such as complementary and sandwich cytoarchitectures, remains a long-lasting challenge. To overcome this challenge, herein we present an acoustic differential bioassembly technique to assemble different cell types at the distinct positions of the acoustic field based on their inherent physical characteristics including cellular size and buoyant density. Specifically, different cell types can be differentially assembled beneath the nodal or the antinode regions of the Faraday wave to form complementary cytoarchitectures, or be selectively positioned at the center or edge area beneath either the nodal or the antinode regions to form sandwich cytoarchitectures. Using this technique, we assemble human induced pluripotent stem cell-derived liver spheroids and endothelial cells into hexagonal cytoarchitecturesin vitroto mimic the cord and sinusoid structures in the hepatic lobules. This hepatic lobule model reconstitutes liver metabolic and synthetic functions, such as albumin secretion and urea production. Overall, the acoustic differential bioassembly technique facilitates the construction of human relevantin vitroorganotypic models with spatially-defined heterocellular architectures, and can potentially find wide applications in tissue engineering and regenerative medicine.

  View details for DOI 10.1088/1758-5090/aca79c

  View details for PubMedID 36541139

• Advanced Point-of-Care Testing Technologies for Human Acute Respiratory Virus Detection. Advanced materials (Deerfield Beach, Fla.) Zhang, Z., Ma, P., Ahmed, R., Wang, J., Akin, D., Soto, F., Liu, B., Li, P., Demirci, U. 2021: e2103646

  Abstract

  The ever-growing global threats to human life caused by the human acute respiratory virus (RV) infections have cost billions of lives, created a significant economic burden, and shaped society for centuries. The timely response to emerging RVs could save human lives and reduce the medical care burden. The development of RV detection technologies is essential for potentially preventing RV pandemic and epidemics. However, commonly used detection technologies lack sensitivity, specificity, and speed, thus often failing to provide the rapid turnaround times. To address this problem, new technologies are devised to address the performance inadequacies of the traditional methods. These emerging technologies offer improvements in convenience, speed, flexibility, and portability of point-of-care test (POCT). Herein, recent developments in POCT are comprehensively reviewed for eight typical acute respiratory viruses. This review discusses the challenges and opportunities of various recognition and detection strategies and discusses these according to their detection principles, including nucleic acid amplification, optical POCT, electrochemistry, lateral flow assays, microfluidics, enzyme-linked immunosorbent assays, and microarrays. The importance of limits of detection, throughput, portability, and specificity when testing clinical samples in resource-limited settings is emphasized. Finally, the evaluation of commercial POCT kits for both essential RV diagnosis and clinical-oriented practices is included.

  View details for DOI 10.1002/adma.202103646

  View details for PubMedID 34623709

• 3DICE coding matrix multidirectional macro-architecture modulates cell organization, shape, and co-cultures endothelization network. Biomaterials Canadas, R. F., Costa, J. B., Mao, Z., Gao, C., Demirci, U., Reis, R. L., Marques, A. P., Oliveira, J. M. 2021; 277: 121112

  Abstract

  Natural extracellular matrix governs cells providing biomechanical and biofunctional outstanding properties, despite being porous and mostly made of soft materials. Among organs, specific tissues present specialized macro-architectures. For instance, hepatic lobules present radial organization, while vascular sinusoids are branched from vertical veins, providing specific biofunctional features. Therefore, it is imperative to mimic such structures while modeling tissues. So far, there is limited capability of coupling oriented macro-structures with interconnected micro-channels in programmable long-range vertical and radial sequential orientations. Herein, a three-directional ice crystal elongation (3DICE) system is presented to code geometries in cryogels. Using 3DICE, guided ice crystals growth templates vertical and radial pores through bulky cryogels. Translucent isotropic and anisotropic architectures of radial or vertical pores are fabricated with tunable mechanical response. Furthermore, 3D combinations of vertical and radial pore orientations are coded at the centimeter scale. Cell morphological response to macro-architectures is demonstrated. The formation of endothelial segments, CYP450 activity, and osteopontin expression, as liver fibrosis biomarkers, present direct response and specific cellular organization within radial, linear, and random architectures. These results unlock the potential of ice-templating demonstrating the relevance of macro-architectures to model tissues, and broad possibilities for drug testing, tissue engineering, and regenerative medicine.

  View details for DOI 10.1016/j.biomaterials.2021.121112

  View details for PubMedID 34488122

• Malignant Ascites in Ovarian Cancer: Cellular, Acellular, and Biophysical Determinants of Molecular Characteristics and Therapy Response. Cancers Rickard, B. P., Conrad, C., Sorrin, A. J., Ruhi, M. K., Reader, J. C., Huang, S. A., Franco, W., Scarcelli, G., Polacheck, W. J., Roque, D. M., Del Carmen, M. G., Huang, H., Demirci, U., Rizvi, I. 2021; 13 (17)

  Abstract

  Ascites refers to the abnormal accumulation of fluid in the peritoneum resulting from an underlying pathology, such as metastatic cancer. Among all cancers, advanced-stage epithelial ovarian cancer is most frequently associated with the production of malignant ascites and is the leading cause of death from gynecologic malignancies. Despite decades of evidence showing that the accumulation of peritoneal fluid portends the poorest outcomes for cancer patients, the role of malignant ascites in promoting metastasis and therapy resistance remains poorly understood. This review summarizes the current understanding of malignant ascites, with a focus on ovarian cancer. The first section provides an overview of heterogeneity in ovarian cancer and the pathophysiology of malignant ascites. Next, analytical methods used to characterize the cellular and acellular components of malignant ascites, as well the role of these components in modulating cell biology, are discussed. The review then provides a perspective on the pressures and forces that tumors are subjected to in the presence of malignant ascites and the impact of physical stress on therapy resistance. Treatment options for malignant ascites, including surgical, pharmacological and photochemical interventions are then discussed to highlight challenges and opportunities at the interface of drug discovery, device development and physical sciences in oncology.

  View details for DOI 10.3390/cancers13174318

  View details for PubMedID 34503128

• Multiparametric biophysical profiling of red blood cells in malaria infection. Communications biology Deshmukh, S. S., Shakya, B., Chen, A., Durmus, N. G., Greenhouse, B., Egan, E. S., Demirci, U. 2021; 4 (1): 697

  Abstract

  Biophysical separation promises label-free, less-invasive methods to manipulate the diverse properties of live cells, such as density, magnetic susceptibility, and morphological characteristics. However, some cellular changes are so minute that they are undetectable by current methods. We developed a multiparametric cell-separation approach to profile cells with simultaneously changing density and magnetic susceptibility. We demonstrated this approach with the natural biophysical phenomenon of Plasmodium falciparum infection, which modifies its host erythrocyte by simultaneously decreasing density and increasing magnetic susceptibility. Current approaches have used these properties separately to isolate later-stage infected cells, but not in combination. We present biophysical separation of infected erythrocytes by balancing gravitational and magnetic forces to differentiate infected cell stages, including early stages for the first time, using magnetic levitation. We quantified height distributions of erythrocyte populations-27 ring-stage synchronized samples and 35 uninfected controls-and quantified their unique biophysical signatures. This platform can thus enable multidimensional biophysical measurements on unique cell types.

  View details for DOI 10.1038/s42003-021-02181-3

  View details for PubMedID 34103669

• Engineering Hydrogel-Based Biomedical Photonics: Design, Fabrication, and Applications. Advanced materials (Deerfield Beach, Fla.) Guimaraes, C. F., Ahmed, R., Marques, A. P., Reis, R. L., Demirci, U. 2021: e2006582

  Abstract

  Light guiding and manipulation in photonics have become ubiquitous in events ranging from everyday communications to complex robotics and nanomedicine. The speed and sensitivity of light-matter interactions offer unprecedented advantages in biomedical optics, data transmission, photomedicine, and detection of multi-scale phenomena. Recently, hydrogels have emerged as a promising candidate for interfacing photonics and bioengineering by combining their light-guiding properties with live tissue compatibility in optical, chemical, physiological, and mechanical dimensions. Herein, the latest progress over hydrogel photonics and its applications in guidance and manipulation of light is reviewed. Physics of guiding light through hydrogels and living tissues, and existing technical challenges in translating these tools into biomedical settings are discussed. Acomprehensive and thorough overview of materials, fabrication protocols, and design architectures used in hydrogel photonics is provided. Finally, recent examples of applying structures such as hydrogel optical fibers, living photonic constructs, and their use as light-driven hydrogel robots, photomedicine tools, and organ-on-a-chip models are described. By providing a critical and selective evaluation of the field's status, this work sets a foundation for the next generation of hydrogel photonic research.

  View details for DOI 10.1002/adma.202006582

  View details for PubMedID 33929771

• Management of COVID-19: Current Status and Future Prospects. Microbes and infection Kabir, M. A., Ahmed, R., Chowdhury, R., Asher Iqbal, S. M., Paulmurugan, R., Demirci, U., Asghar, W. 2021: 104832

  Abstract

  COVID-19, a highly transmissible pandemic disease, affecting millions of lives around the world. Severely infected patients show acute respiratory distress symptoms. Sustainable management strategies are required to save the lives of the infected people and further preventing spread of the virus. Diagnosis, treatment, and vaccination development initiatives are already exhibited from the scientific community to fight against this virus. In this review, we primarily discuss the management strategies including Prevention of spread, prophylaxis, vaccinations, and treatment for COVID-19. Further, analysis of vaccine development status and performance are also briefly discussed. Global social and economic impact of COVID-19 are also analyzed as part of this review.

  View details for DOI 10.1016/j.micinf.2021.104832

  View details for PubMedID 33872807

• Towards Microfluidic-Based Exosome Isolation and Detection for Tumor Therapy. Nano today Wang, J., Ma, P., Kim, D. H., Liu, B., Demirci, U. 2021; 37

  Abstract

  Exosomes are a class of cell-secreted, nano-sized extracellular vesicles with a bilayer membrane structure of 30-150 nm in diameter. Their discovery and application have brought breakthroughs in numerous areas, such as liquid biopsies, cancer biology, drug delivery, immunotherapy, tissue repair, and cardiovascular diseases. Isolation of exosomes is the first step in exosome-related research and its applications. Standard benchtop exosome separation and sensing techniques are tedious and challenging, as they require large sample volumes, multi-step operations that are complex and time-consuming, requiring cumbersome and expensive instruments. In contrast, microfluidic platforms have the potential to overcome some of these limitations, owing to their high-precision processing, ability to handle liquids at a microscale, and integrability with various functional units, such as mixers, actuators, reactors, separators, and sensors. These platforms can optimize the detection process on a single device, representing a robust and versatile technique for exosome separation and sensing to attain high purity and high recovery rates with a short processing time. Herein, we overview microfluidic strategies for exosome isolation based on their hydrodynamic properties, size filtration, acoustic fields, immunoaffinity, and dielectrophoretic properties. We focus especially on advances in label-free isolation of exosomes with active biological properties and intact morphological structures. Further, we introduce microfluidic techniques for the detection of exosomal proteins and RNAs with high sensitivity, high specificity, and low detection limits. We summarize the biomedical applications of exosome-mediated therapeutic delivery targeting cancer cells. To highlight the advantages of microfluidic platforms, conventional techniques are included for comparison. Future challenges and prospects of microfluidics towards exosome isolation applications are also discussed. Although the use of exosomes in clinical applications still faces biological, technical, regulatory, and market challenges, in the foreseeable future, recent developments in microfluidic technologies are expected to pave the way for tailoring exosome-related applications in precision medicine.

  View details for DOI 10.1016/j.nantod.2020.101066

  View details for PubMedID 33777166

• Reversible Design of Dynamic Assemblies at Small Scales ADVANCED INTELLIGENT SYSTEMS Soto, F., Wang, J., Deshmukh, S., Demirci, U. 2021; 3 (4)

  View details for DOI 10.1002/aisy.202000193

  View details for Web of Science ID 000669806800008

• Reversible Design of Dynamic Assemblies at Small Scales. Advanced intelligent systems (Weinheim an der Bergstrasse, Germany) Soto, F., Wang, J., Deshmukh, S., Demirci, U. 2021; 3 (4)

  Abstract

  Emerging bottom-up fabrication methods have enabled the assembly of synthetic colloids, microrobots, living cells, and organoids to create intricate structures with unique properties that transcend their individual components. This review provides an access point to the latest developments in externally driven assembly of synthetic and biological components. In particular, we emphasize reversibility, which enables the fabrication of multiscale systems that would not be possible under traditional techniques. Magnetic, acoustic, optical, and electric fields are the most promising methods for controlling the reversible assembly of biological and synthetic subunits since they can reprogram their assembly by switching on/off the external field or shaping these fields. We feature capabilities to dynamically actuate the assembly configuration by modulating the properties of the external stimuli, including frequency and amplitude. We describe the design principles which enable the assembly of reconfigurable structures. Finally, we foresee that the high degree of control capabilities offered by externally driven assembly will enable broad access to increasingly robust design principles towards building advanced dynamic intelligent systems.

  View details for DOI 10.1002/aisy.202000193

  View details for PubMedID 35663639

  View details for PubMedCentralID PMC9165726

• Diagnosis For COVID-19: Current Status and Future Prospects. Expert review of molecular diagnostics Kabir, A., Ahmed, R., Iqbal, S. M., Chowdhury, R., Paulmurugan, R., Demirci, U., Asghar, W. 2021

  Abstract

  INTRODUCTION: Coronavirus disease 2019 (COVID-19), a respiratory illness caused by novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), had its first detection in December 2019 in Wuhan (China) and spread across the world. In March 2020, the World Health Organization (WHO) declared COVID-19 a pandemic disease. The utilization of prompt and accurate molecular diagnosis of SARS-CoV-2 virus, isolating the infected patients, and treating them are the keys to managing this unprecedented pandemic. International travel acted as a catalyst for the widespread transmission of the virus.Areas Covered:This review discusses phenotype, structural, and molecular evolution of recognition elements and primers, its detection in the laboratory, and at point of care. Further, market analysis of commercial products and their performance is also evaluated, providing new ways to confront the ongoing global public health emergency.Expert Commentary:The outbreak for COVID-19 created mammoth chaos to the healthcare sector, and still, day by day, new epicenters for the outbreak are being reported. Emphasis should be placed on developing more effective, rapid, and early diagnostic devices. The testing laboratories should invest more in clinically relevant multiplexed and scalable detection tools to fight against a pandemic like this where massive demand for testing exists.

  View details for DOI 10.1080/14737159.2021.1894930

  View details for PubMedID 33621145

• Micropatterned Surfaces Expose the Coupling between Actin Cytoskeleton-Lamin/Nesprin and Nuclear Deformability of Breast Cancer Cells with Different Malignancies. Advanced biology Antmen, E. n., Demirci, U. n., Hasirci, V. n. 2021; 5 (1): e2000048

  Abstract

  Mechanotransduction proteins transfer mechanical stimuli through nucleo-cytoskeletal coupling and affect the nuclear morphology of cancer cells. However, the contribution of actin filament integrity has never been studied directly. It is hypothesized that differences in nuclear deformability of cancer cells are influenced by the integrity of actin filaments. In this study, transparent micropatterned surfaces as simple tools to screen cytoskeletal and nuclear distortions are presented. Surfaces decorated with micropillars are used to culture and image breast cancer cells and quantify their deformation using shape descriptors (circularity, area, perimeter). Using two drugs (cytochalasin D and jasplakinolide), actin filaments are disrupted. Deformation of cells on micropillars is decreased upon drug treatment as shown by increased circularity. However, the effect is much smaller on benign MCF10A than on malignant MCF7 and MDAMB231 cells. On micropatterned surfaces, molecular analysis shows that Lamin A/C and Nesprin-2 expressions decreased but, after drug treatment, increased in malignant cells but not in benign cells. These findings suggest that Lamin A/C, Nesprin-2 and actin filaments are critical in mechanotransduction of cancer cells. Consequently, transparent micropatterned surfaces can be used as image analysis platforms to provide robust, high throughput measurements of nuclear deformability of cancer cells, including the effect of cytoskeletal elements.

  View details for DOI 10.1002/adbi.202000048

  View details for PubMedID 33724728

• Mitochondria-Rich Extracellular Vesicles Rescue Patient-Specific Cardiomyocytes From Doxorubicin Injury: Insights Into the SENECA Trial. JACC. CardioOncology O'Brien, C. G., Ozen, M. O., Ikeda, G., Vaskova, E., Jung, J. H., Bayardo, N., Santoso, M. R., Shi, L., Wahlquist, C., Jiang, Z., Jung, Y., Zeng, Y., Egan, E., Sinclair, R., Gee, A., Witteles, R., Mercola, M., Svensson, K. J., Demirci, U., Yang, P. C. 2021; 3 (3): 428-440

  Abstract

  Anthracycline-induced cardiomyopathy (AIC) is a significant source of morbidity and mortality in cancer survivors. The role of mesenchymal stem cells (MSCs) in treating AIC was evaluated in the SENECA trial, a Phase 1 National Heart, Lung, and Blood Institute-sponsored study, but the mechanisms underpinning efficacy in human tissue need clarification.The purpose of this study was to perform an in vitro clinical trial evaluating the efficacy and putative mechanisms of SENECA trial-specific MSCs in treating doxorubicin (DOX) injury, using patient-specific induced pluripotent stem cell-derived cardiomyocytes (iCMs) generated from SENECA patients.Patient-specific iCMs were injured with 1 μmol/L DOX for 24 hours, treated with extracellular vesicles (EVs) from MSCs by either coculture or direct incubation and then assessed for viability and markers of improved cellular physiology. MSC-derived EVs were separated into large extracellular vesicles (L-EVs) (>200 nm) and small EVs (<220nm) using a novel filtration system.iCMs cocultured with MSCs in a transwell system demonstrated improved iCM viability and attenuated apoptosis. L-EVs but not small EVs recapitulated this therapeutic effect. L-EVs were found to be enriched in mitochondria, which were shown to be taken up by iCMs. iCMs treated with L-EVs demonstrated improved contractility, reactive oxygen species production, ATP production, and mitochondrial biogenesis. Inhibiting L-EV mitochondrial function with 1-methyl-4-phenylpyridinium attenuated efficacy.L-EV-mediated mitochondrial transfer mitigates DOX injury in patient-specific iCMs. Although SENECA was not designed to test MSC efficacy, consistent tendencies toward a positive effect were observed across endpoints. Our results suggest a mechanism by which MSCs may improve cardiovascular performance in AIC independent of regeneration, which could inform future trial design evaluating the therapeutic potential of MSCs.

  View details for DOI 10.1016/j.jaccao.2021.05.006

  View details for PubMedID 34604804

  View details for PubMedCentralID PMC8463733

• Wearable Collector for Noninvasive Sampling of SARS-CoV-2 from Exhaled Breath for Rapid Detection. ACS applied materials & interfaces Soto, F., Ozen, M. O., Guimarães, C. F., Wang, J., Hokanson, K., Ahmed, R., Reis, R. L., Paulmurugan, R., Demirci, U. 2021

  Abstract

  Airborne transmission of exhaled virus can rapidly spread, thereby increasing disease progression from local incidents to pandemics. Due to the COVID-19 pandemic, states and local governments have enforced the use of protective masks in public and work areas to minimize the disease spread. Here, we have leveraged the function of protective face coverings toward COVID-19 diagnosis. We developed a user-friendly, affordable, and wearable collector. This noninvasive platform is integrated into protective masks toward collecting airborne virus in the exhaled breath over the wearing period. A viral sample was sprayed into the collector to model airborne dispersion, and then the enriched pathogen was extracted from the collector for further analytical evaluation. To validate this design, qualitative colorimetric loop-mediated isothermal amplification, quantitative reverse transcription polymerase chain reaction, and antibody-based dot blot assays were performed to detect the presence of SARS-CoV-2. We envision that this platform will facilitate sampling of current SARS-CoV-2 and is potentially broadly applicable to other airborne diseases for future emerging pandemics.

  View details for DOI 10.1021/acsami.1c09309

  View details for PubMedID 34428374

• Engineering Polysaccharide-Based Hydrogel Photonic Constructs: From Multiscale Detection to the Biofabrication of Living Optical Fibers. Advanced materials (Deerfield Beach, Fla.) Guimarães, C. F., Ahmed, R., Mataji-Kojouri, A., Soto, F., Wang, J., Liu, S., Stoyanova, T., Marques, A. P., Reis, R. L., Demirci, U. 2021: e2105361

  Abstract

  Solid-state optics has been the pillar of modern digital age. Integrating soft hydrogel materials with micro/nanooptics could expand the horizons of photonics for bioengineering. Here, wet-spun multilayer hydrogel fibers are engineered through ionic-crosslinked natural polysaccharides that serve as multifunctional platforms. The resulting flexible hydrogel structure and reversible crosslinking provide tunable design properties such as adjustable refractive index and fusion splicing. Modulation of the optical readout via physical stimuli, including shape, compression, and multiple optical inputs/outputs is demonstrated. The unique permeability of the hydrogels is also combined with plasmonic nanoparticles for molecular detection of SARS-CoV-2 in fiber-coupled biomedical swabs. A tricoaxial 3D printing nozzle is then employed for the continuous fabrication of living optical fibers. Light interaction with living cells enables the quantification and digitalization of complex biological phenomena such as 3D cancer progression and drug susceptibility. These fibers pave the way for advances in biomaterial-based photonics and biosensing platforms.

  View details for DOI 10.1002/adma.202105361

  View details for PubMedID 34617338

• Progress and challenges in biomarker enrichment for cancer early detection Progress in Biomedical Engineering Sinawang, P., Soto, F., Ozen, M. O., Akin, D., Demirci, U. 2021; 3 (4)

  View details for DOI 10.1088/2516-1091/ac1ea3

• Emerging biofabrication approaches for gastrointestinal organoids towards patient specific cancer models. Cancer letters Soto, F. n., Guimarães, C. F., Reis, R. L., Franco, W. n., Rizvi, I. n., Demirci, U. n. 2021

  Abstract

  Tissue engineered organoids are simple biomodels that can emulate the structural and functional complexity of specific organs. Here, we review developments in three-dimensional (3D) artificial cell constructs to model gastrointestinal dynamics towards cancer diagnosis. We describe bottom-up approaches to fabricate close-packed cell aggregates, from the use of biochemical and physical cues to guide the self-assembly of organoids, to the use of engineering approaches, including 3D printing/additive manufacturing and external field-driven protocols. Finally, we outline the main challenges and possible risks regarding the potential translation of gastrointestinal organoids from laboratory settings to patient-specific models in clinical applications.

  View details for DOI 10.1016/j.canlet.2021.01.023

  View details for PubMedID 33577978

• Engineering the Interaction Dynamics between Nano-Topographical Immunocyte-Templated Micromotors across Scales from Ions to Cells. Small (Weinheim an der Bergstrasse, Germany) Wang, J., Ahmed, R., Zeng, Y., Fu, K., Soto, F., Sinclair, B., Soh, H. T., Demirci, U. 2020: e2005185

  Abstract

  Manufacturing mobile artificial micromotors with structural design factors, such as morphology nanoroughness and surface chemistry, can improve the capture efficiency through enhancing contact interactions with their surrounding targets. Understanding the interplay of such parameters targeting high locomotion performance and high capture efficiency at the same time is of paramount importance, yet, has so far been overlooked. Here, an immunocyte-templated nano-topographical micromotor is engineered and their interactions with various targets across multiple scales, from ions to cells are investigated. The macrophage templated nanorough micromotor demonstrates significantly increased surface interactions and significantly improved and highly efficient removal of targets from complex aqueous solutions, including in plasma and diluted blood, when compared to smooth synthetic material templated micromotors with the same size and surface chemistry. These results suggest that the surface nanoroughness of the micromotors for the locomotion performance and interactions with the multiscale targets should be considered simultaneously, for they are highly interconnected in design considerations impacting applications across scales.

  View details for DOI 10.1002/smll.202005185

  View details for PubMedID 33174334

• Atlas of Exosomal microRNAs Secreted From Human iPSC-Derived Cardiac Cell Types. Circulation Chandy, M., Rhee, J., Ozen, M. O., Williams, D. R., Pepic, L., Liu, C., Zhang, H., Malisa, J., Lau, E., Demirci, U., Wu, J. C. 2020; 142 (18): 1794–96

  View details for DOI 10.1161/CIRCULATIONAHA.120.048364

  View details for PubMedID 33136510

• Enhancing the nanoplasmonic signal by a nanoparticle sandwiching strategy to detect viruses APPLIED MATERIALS TODAY Inci, F., Karaaslan, M., Mataji-Kojouri, A., Shah, P., Saylan, Y., Zeng, Y., Avadhani, A., Sinclair, R., Lau, D., Demirci, U. 2020; 20

  View details for DOI 10.1016/j.apmt.2020.100709

  View details for Web of Science ID 000598351900006

• Magnetic levitational bioassembly of 3D tissue construct in space. Science advances Parfenov, V. A., Khesuani, Y. D., Petrov, S. V., Karalkin, P. A., Koudan, E. V., Nezhurina, E. K., Pereira, F. D., Krokhmal, A. A., Gryadunova, A. A., Bulanova, E. A., Vakhrushev, I. V., Babichenko, I. I., Kasyanov, V., Petrov, O. F., Vasiliev, M. M., Brakke, K., Belousov, S. I., Grigoriev, T. E., Osidak, E. O., Rossiyskaya, E. I., Buravkova, L. B., Kononenko, O. D., Demirci, U., Mironov, V. A. 2020; 6 (29): eaba4174

  Abstract

  Magnetic levitational bioassembly of three-dimensional (3D) tissue constructs represents a rapidly emerging scaffold- and label-free approach and alternative conceptual advance in tissue engineering. The magnetic bioassembler has been designed, developed, and certified for life space research. To the best of our knowledge, 3D tissue constructs have been biofabricated for the first time in space under microgravity from tissue spheroids consisting of human chondrocytes. Bioassembly and sequential tissue spheroid fusion presented a good agreement with developed predictive mathematical models and computer simulations. Tissue constructs demonstrated good viability and advanced stages of tissue spheroid fusion process. Thus, our data strongly suggest that scaffold-free formative biofabrication using magnetic fields is a feasible alternative to traditional scaffold-based approaches, hinting a new perspective avenue of research that could significantly advance tissue engineering. Magnetic levitational bioassembly in space can also advance space life science and space regenerative medicine.

  View details for DOI 10.1126/sciadv.aba4174

  View details for PubMedID 32743068

• A confirmatory test for sperm in sexual assault samples using a microfluidic-integrated cell phone imaging system. Forensic science international. Genetics Deshmukh, S., Inci, F., Karaaslan, M. G., Ogut, M. G., Duncan, D., Klevan, L., Duncan, G., Demirci, U. 2020; 48: 102313

  Abstract

  Rapid and efficient processing of sexual assault evidence to accelerate forensic investigation and decrease casework backlogs is urgently needed. Therefore, the standardized protocols currently used in forensic laboratories can benefit from continued innovation to handle the increasing number and complexity of samples being submitted to forensic labs. To our knowledge, there is currently no available rapid and portable forensic screening technology based on a confirmatory test for sperm identification in a sexual assault kit. Here, we present a novel forensic sample screening tool, i.e., a microchip integrated with a portable cell phone imaging platform that records and processes images for further investigation and storage. The platform (i) precisely and rapidly screens swab samples (<15 min after sample preparation on-chip); (ii) selectively captures sperm from mock sexual assault samples using a novel and previously published SLeX-based surface chemistry treatment (iii) separates non-sperm contents (epithelial cells and debris in this case) out of the channel by flow prior to imaging; (iv) captures cell phone images on a portable cellphone-integrated imaging platform, (v) quantitatively differentiates sperm cells from epithelial cells, using a morphology detection code that leverages Laplacian of Gaussian and Hough gradient transform methods; (vi) is sensitive within a forensic cut-off (>95% accuracy) compared to the manual counts; (vii) provides a cost-effective and timely solution to a problem which in the past has taken a great deal of time; and (viii) handles small volumes of sample (20 muL). This integration of the cellphone imaging platform and cell recognition algorithms with disposable microchips can be a new direction toward a direct visual test to screen and differentiate sperm from epithelial cell types in forensic samples for a crime laboratory scenario. With further development, this integrated platform could assist a sexual assault nurse examiner (SANE) in a hospital or sexual assault treatment center facility to flag sperm-containing samples prior to further downstream testing.

  View details for DOI 10.1016/j.fsigen.2020.102313

  View details for PubMedID 32570000

• Enhancing cell packing in buckyballs by acoustofluidic activation. Biofabrication Ren, T., Steiger, W., Chen, P., Ovsianikov, A., Demirci, U. 2020; 12 (2): 025033

  Abstract

  How to pack materials into well-defined volumes efficiently has been a longstanding question of interest to physicists, material scientists, and mathematicians as these materials have broad applications ranging from shipping goods in commerce to seeds in agriculture and to spheroids in tissue engineering. How many marbles or gumball candies can you pack into a jar?Although these seem to be idle questions they have been studied for centuries and have recently become of greater interest with their broadening applications in science and medicine. Here, we study a similar problem where we try to pack cells into a spherical porous buckyball structure. The experimental limitations are short of the theoretical maximum packing density due to the microscale of the structures that the cells are being packed into. We show that we can pack more cells into a confined micro-structure (buckyball cage) by employing acoustofluidic activation and their hydrodynamic effect at the bottom of a liquid-carrier chamber compared to randomly dropping cells onto these buckyballs by gravity. Although, in essence, cells would be expected to achieve a higher maximum volume fraction than marbles in a jar, given that they can squeeze and reshape and reorient their structure, the packing density of cells into the spherical buckyball cages are far from this theoretical limit. This is mainly dictated by the experimental limitations of cells washing away as well as being loaded into the chamber.

  View details for DOI 10.1088/1758-5090/ab76d9

  View details for PubMedID 32229710

• Flow-induced Shear Stress Confers Resistance to Carboplatin in an Adherent Three-Dimensional Model for Ovarian Cancer: A Role for EGFR-Targeted Photoimmunotherapy Informed by Physical Stress. Journal of clinical medicine Nath, S., Pigula, M., Khan, A. P., Hanna, W., Ruhi, M. K., Dehkordy, F. M., Pushpavanam, K., Rege, K., Moore, K., Tsujita, Y., Conrad, C., Inci, F., Carmen, M. G., Franco, W., Celli, J. P., Demirci, U., Hasan, T., Huang, H., Rizvi, I. 2020; 9 (4)

  Abstract

  A key reason for the persistently grim statistics associated with metastatic ovarian cancer is resistance to conventional agents, including platinum-based chemotherapies. A major source of treatment failure is the high degree of genetic and molecular heterogeneity, which results from significant underlying genomic instability, as well as stromal and physical cues in the microenvironment. Ovarian cancer commonly disseminates via transcoelomic routes to distant sites, which is associated with the frequent production of malignant ascites, as well as the poorest prognosis. In addition to providing a cell and protein-rich environment for cancer growth and progression, ascitic fluid also confers physical stress on tumors. An understudied area in ovarian cancer research is the impact of fluid shear stress on treatment failure. Here, we investigate the effect of fluid shear stress on response to platinum-based chemotherapy and the modulation of molecular pathways associated with aggressive disease in a perfusion model for adherent 3D ovarian cancer nodules. Resistance to carboplatin is observed under flow with a concomitant increase in the expression and activation of the epidermal growth factor receptor (EGFR) as well as downstream signaling members mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) and extracellular signal-regulated kinase (ERK). The uptake of platinum by the 3D ovarian cancer nodules was significantly higher in flow cultures compared to static cultures. A downregulation of phospho-focal adhesion kinase (p-FAK), vinculin, and phospho-paxillin was observed following carboplatin treatment in both flow and static cultures. Interestingly, low-dose anti-EGFR photoimmunotherapy (PIT), a targeted photochemical modality, was found to be equally effective in ovarian tumors grown under flow and static conditions. These findings highlight the need to further develop PIT-based combinations that target the EGFR, and sensitize ovarian cancers to chemotherapy in the context of flow-induced shear stress.

  View details for DOI 10.3390/jcm9040924

  View details for PubMedID 32231055

• Tunable Fano-Resonant Metasurfaces on a Disposable Plastic-Template for Multimodal and Multiplex Biosensing. Advanced materials (Deerfield Beach, Fla.) Ahmed, R., Ozen, M. O., Karaaslan, M. G., Prator, C. A., Thanh, C., Kumar, S., Torres, L., Iyer, N., Munter, S., Southern, S., Henrich, T. J., Inci, F., Demirci, U. 2020: e1907160

  Abstract

  Metasurfaces are engineered nanostructured interfaces that extend the photonic behavior of natural materials, and they spur many breakthroughs in multiple fields, including quantum optics, optoelectronics, and biosensing. Recent advances in metasurface nanofabrication enable precise manipulation of light-matter interactions at subwavelength scales. However, current fabrication methods are costly and time-consuming and have a small active area with low reproducibility due to limitations in lithography, where sensing nanosized rare biotargets requires a wide active surface area for efficient binding and detection. Here, a plastic-templated tunable metasurface with a large active area and periodic metal-dielectric layers to excite plasmonic Fano resonance transitions providing multimodal and multiplex sensing of small biotargets, such as proteins and viruses, is introduced. The tunable Fano resonance feature of the metasurface is enabled via chemical etching steps to manage nanoperiodicity of the plastic template decorated with plasmonic layers and surrounding dielectric medium. This metasurface integrated with microfluidics further enhances the light-matter interactions over a wide sensing area, extending data collection from 3D to 4D by tracking real-time biomolecular binding events. Overall, this work resolves cost- and complexity-related large-scale fabrication challenges and improves multilayer sensitivity of detection in biosensing applications.

  View details for DOI 10.1002/adma.201907160

  View details for PubMedID 32201997

• Multiscale brain research on a microfluidic chip. Lab on a chip Zhao, Y., Demirci, U., Chen, Y., Chen, P. 2020

  Abstract

  One major challenge in current brain research is generating an integrative understanding of the brain's functions and disorders from its multiscale neuronal architectures and connectivity. Thus, innovative neurotechnology tools are urgently required for deciphering the multiscale functional and structural organizations of the brain at hierarchical scales from the molecular to the organismal level by multiple brain research initiatives launched by the European Union, United States, Australia, Canada, China, Korea, and Japan. To meet this demand, microfluidic chips (muFCs) have rapidly evolved as a trans-scale neurotechnological toolset to enable multiscale studies of the brain due to their unique advantages in flexible microstructure design, multifunctional integration, accurate microenvironment control, and capacity for automatic sample processing. Here, we review the recent progress in applying innovative muFC-based neuro-technologies to promote multiscale brain research and uniquely focus on representative applications of muFCs to address challenges in brain research at each hierarchical level. We discuss the current trend of combinational applications of muFCs with other neuro- and biotechnologies, including optogenetics, brain organoids, and 3D bioprinting, for better multiscale brain research. In addition, we offer our insights into the existing outstanding questions at each hierarchical level of brain research that could potentially be addressed by advancing microfluidic techniques. This review will serve as a timely guide for bioengineers and neuroscientists to develop and apply muFC-based neuro-technologies for promoting basic and translational brain research.

  View details for DOI 10.1039/c9lc01010f

  View details for PubMedID 32150176

• Fabrication of calcium phosphate 3D scaffolds for bone repair using magnetic levitational assembly. Scientific reports Parfenov, V. A., Mironov, V. A., Koudan, E. V., Nezhurina, E. K., Karalkin, P. A., Pereira, F. D., Petrov, S. V., Krokhmal, A. A., Aydemir, T., Vakhrushev, I. V., Zobkov, Y. V., Smirnov, I. V., Fedotov, A. Y., Demirci, U., Khesuani, Y. D., Komlev, V. S. 2020; 10 (1): 4013

  Abstract

  The calcium phosphate particles can be used as building blocks for fabrication of 3D scaffolds intended for bone tissue engineering. This work presents for the first time a rapid creation of 3D scaffolds using magnetic levitation of calcium phosphate particles. Namely, tricalcium phosphate particles of equal size and certain porosity are used, which undergo the process of recrystallization after magnetic levitational assembly of the scaffold to ensure stitching of the scaffold. Label-free levitational assembly is achieved by using a custom-designed magnetic system in the presence of gadolinium salts, which allows the levitation of calcium phosphate particles. Chemical transformation of tricalcium- to octacalcium phosphate under the condition of magnetic levitation in non-homogeneous magnetic field is also demonstrated. This approach allows obtaining rapidly the octacalcium phosphate phase in the final 3D product, which is biocompatible.

  View details for DOI 10.1038/s41598-020-61066-3

  View details for PubMedID 32132636

• A disposable microfluidic-integrated hand-held plasmonic platform for protein detection APPLIED MATERIALS TODAY Inci, F., Saylan, Y., Kojouri, A., Ogut, M., Denizli, A., Demirci, U. 2020; 18

  View details for DOI 10.1016/j.apmt.2019.100478

  View details for Web of Science ID 000530652300026

• Total Microfluidic chip for Multiplexed diagnostics (ToMMx). Biosensors & bioelectronics Ozen, M. O., Sridhar, K., Ogut, M. G., Shanmugam, A., Avadhani, A. S., Kobayashi, Y., Wu, J. C., Haddad, F., Demirci, U. 2020; 150: 111930

  Abstract

  Microfluidic technologies offer new platforms for biosensing in various clinical and point-of-care (POC) applications. Currently, at the clinical settings, the gold standard diagnostic platforms for multiplexed sensing are multi-step, time consuming, requiring expensive and bulky instruments with a constant need of electricity which makes them unsuitable for resource-limited or POC settings. These technologies are often limited by logistics, costly assays and regular maintenance. Although there have been several attempts to miniaturize these diagnostic platforms, they stand short of batch fabrication and they are dependent on complementary components such as syringe pumps. Here, we demonstrated the development and clinical testing of a disposable, multiplexed sensing device (ToMMx), which is a portable, high-throughput and user-friendly microfluidic platform. It was built with inexpensive plastic materials and operated manually without requiring electrical power and extensive training. We validated this platform in a small cohort of 50 clinical samples from patients with cardiovascular diseases and healthy controls. The platform is rapid and gives quantifiable results with high sensitivity, as low as 5.29pg/mL, from only a small sample volume (4muL). ToMMx platform was compared side-by-side with commercial ELISA kits where the total assay time is reduced 15-fold, from 5h to 20min. This technology platform is broadly applicable to various diseases with well-known biomarkers in diagnostics and monitoring, especially with potential future impact at the POC settings.

  View details for DOI 10.1016/j.bios.2019.111930

  View details for PubMedID 31929083

• A Circulating Bioreactor Reprograms Cancer Cells Toward a More Mesenchymal Niche. Advanced biosystems Calamak, S., Ermis, M., Sun, H., Islam, S., Sikora, M., Nguyen, M., Hasirci, V., Steinmetz, L. M., Demirci, U. 2020; 4 (2): e1900139

  Abstract

  Cancer is a complex and heterogeneous disease, and cancer cells dynamically interact with the mechanical microenvironment such as hydrostatic pressure, fluid shear, and interstitial flow. These factors play an essential role in cell fate and circulating tumor cell heterogeneity, and can influence the cellular phenotype. In this study, a peristaltic continuous flow reactor is designed and applied to HCT-116 colorectal carcinoma cells to mimic the fluid dynamics of circulation. With this intervention, a CD44/CD24-cell subpopulation emerges, and 100 genes are significantly regulated. The expression of cells at 4 h in the flow reactor is very similar to TGF-ß treatment, which is an inducer of epithelial-mesenchymal transition. ATF3 and SERPINE1 are significantly upregulated in these groups, suggesting that the mesenchymal transition is induced through this signaling pathway. This flow reactor model is satisfactory on its own to reprogram colorectal cancer cells toward a more mesenchymal niche mimicking circulation of the blood.

  View details for DOI 10.1002/adbi.201900139

  View details for PubMedID 32293132

• A Circulating Bioreactor Reprograms Cancer Cells Toward a More Mesenchymal Niche ADVANCED BIOSYSTEMS Calamak, S., Ermis, M., Sun, H., Islam, S., Sikora, M., Nguyen, M., Hasirci, V., Steinmetz, L. M., Demirci, U. 2020

  View details for DOI 10.1002/adbi.201900139

  View details for Web of Science ID 000506961500001

• Entangled Nanoplasmonic Cavities for Estimating Thickness of Surface-Adsorbed Layers. ACS nano Mataji-Kojouri, A. n., Ozen, M. O., Shahabadi, M. n., Inci, F. n., Demirci, U. n. 2020

  Abstract

  Plasmonic sensors provide real-time and label-free detection of biotargets with unprecedented sensitivity and detection limit. However, they usually lack the ability to estimate the thickness of the target layer formed on top of the sensing surface. Here, we report a sensing modality based on reflection spectroscopy of a nanoplasmonic Fabry-Perot cavity array, which exhibits characteristics of both surface plasmon polaritons and localized plasmon resonances and outperforms its conventional counterparts by providing the thickness of the surface-adsorbed layers. Through numerical simulations, we demonstrate that the designed plasmonic surface resembles two entangled Fabry-Perot cavities excited from both ends. Performance of the device is evaluated by studying sensor response in the refractive index (RI) measurement of aqueous glycerol solutions and during formation of a surface-adsorbed layer consisting of protein (i.e., NeutrAvidin) molecules. By tracking the resonance wavelengths of the two modes of the nanoplasmonic surface, it is therefore possible to measure the thickness of a homogeneous adsorbed layer and RI of the background solution with precisions better than 4 nm and 0.0001 RI units. Using numerical simulations, we show that the thickness estimation algorithm can be extended for layers consisting of nanometric analytes adsorbed on an antibody-coated sensor surface. Furthermore, performance of the device has been evaluated to detect exosomes. By providing a thickness estimation for adsorbed layers and differentiating binding events from background RI variations, this device can potentially supersede conventional plasmonic sensors.

  View details for DOI 10.1021/acsnano.0c02797

  View details for PubMedID 32639713

• The Paracrine Function of Mesenchymal Stem Cells in Response to Pulsed Focused Ultrasound CELL TRANSPLANTATION Razavi, M., Rezaee, M., Telichko, A., Inan, H., Dahl, J., Demirci, U., Thakor, A. S. 2020; 29: 963689720965478

  Abstract

  We studied the paracrine function of mesenchymal stem cells (MSCs) derived from various sources in response to pulsed focused ultrasound (pFUS). Human adipose tissue (AD), bone marrow (BM), and umbilical cord (UC) derived MSCs were exposed to pFUS at two intensities: 0.45 W/cm2 ISATA (310 kPa PNP) and 1.3 W/cm2 ISATA (540 kPa PNP). Following pFUS, the viability and proliferation of MSCs were assessed using a hemocytometer and confocal microscopy, and their secreted cytokine profile determined using a multiplex ELISA. Our findings showed that pFUS can stimulate the production of immunomodulatory, anti-inflammatory, and angiogenic cytokines from MSCs which was dependent on both the source of MSC being studied and the acoustic intensity employed. These important findings set the foundation for additional mechanistic and validation studies using this novel noninvasive and clinically translatable technology for modulating MSC biology.

  View details for DOI 10.1177/0963689720965478

  View details for Web of Science ID 000606584100044

  View details for PubMedID 33028105

  View details for PubMedCentralID PMC7784560

• A bioartificial liver support system integrated with a DLM/GelMA-based bioengineered whole liver for prevention of hepatic encephalopathy via enhanced ammonia reduction. Biomaterials science Wu, G. n., Wu, D. n., Lo, J. n., Wang, Y. n., Wu, J. n., Lu, S. n., Xu, H. n., Zhao, X. n., He, Y. n., Li, J. n., Demirci, U. n., Wang, S. n. 2020

  Abstract

  Although bioartificial liver support systems (BLSSs) play an essential role in maintaining partial liver functions and detoxification for liver failure patients, hepatocytes are unanimously seeded in biomaterials, which lack the hierarchal structures and mechanical cues of native liver tissues. To address this challenge, we developed a new BLSS by combining a decellularized liver matrix (DLM)/GelMA-based bioengineered whole liver and a perfusion-based, oxygenated bioreactor. The novel bioengineered whole liver was fabricated by integrating photocrosslinkable gelatin (GelMA) and hepatocytes into a DLM. The combination of GelMA and the DLM not only provided a biomimetic extracellular microenvironment (ECM) for enhanced cell immobilization and growth with elevated hepatic functions (e.g., albumin secretion and CYP activities), but also provided biomechanical support to maintain the native structure of the liver. In addition, the perfusion-based, oxygenated bioreactor helped deliver oxygen to the interior tissues of the bioengineered liver, which was of importance for long-term culture. Most importantly, this new bioengineered whole liver decreased ammonia concentration by 45%, whereas direct seeding of hepatocytes in a naked DLM showed no significant reduction. Thus, the developed BLSS integrated with the DLM/GelMA-based bioengineered whole liver can potentially help elevate liver functions and prevent HE in liver failure patients while waiting for liver transplantation.

  View details for DOI 10.1039/c9bm01879d

  View details for PubMedID 32307491

• Label-free imaging of exosomes using depth scanning correlation (DSC) interferometric microscopy Aygun, U., Ozkumur, A., Durmus, N., Demirci, U., Urey, H., Shaked, N. T., Hayden, O. SPIE-INT SOC OPTICAL ENGINEERING. 2020

  View details for DOI 10.1117/12.2543242

  View details for Web of Science ID 000558221300008

• Levitating Cells to Sort the Fit and the Fat. Advanced biosystems Puluca, N. n., Durmus, N. G., Lee, S. n., Belbachir, N. n., Galdos, F. X., Ogut, M. G., Gupta, R. n., Hirano, K. I., Krane, M. n., Lange, R. n., Wu, J. C., Wu, S. M., Demirci, U. n. 2020: e1900300

  Abstract

  Density is a core material property and varies between different cell types, mainly based on differences in their lipid content. Sorting based on density enables various biomedical applications such as multi-omics in precision medicine and regenerative repair in medicine. However, a significant challenge is sorting cells of the same type based on density differences. Here, a new method for real-time monitoring and sorting of single cells based on their inherent levitation profiles driven by their lipid content is reported. As a model system, human-induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs) from a patient with neutral lipid storage disease (NLSD) due to loss of function of adipose triglyceride lipase (ATGL) resulting in abnormal lipid storage in cardiac muscle are used. This levitation-based strategy detects subpopulations within ATGL-deficient hiPSC-CMs with heterogenous lipid content, equilibrating at different levitation heights due to small density differences. In addition, sorting of these differentially levitating subpopulations are monitored in real time. Using this approach, sorted healthy and diseased hiPSC-CMs maintain viability and function. Pixel-tracking technologies show differences in contraction between NLSD and healthy hiPSC-CMs. Overall, this is a unique approach to separate diseased cell populations based on their intracellular lipid content that cannot be achieved using traditional flow cytometry techniques.

  View details for DOI 10.1002/adbi.201900300

  View details for PubMedID 32352239

• BIO-INSPIRED MAGNETIC BEADS FOR ISOLATION OF SPERM FROM HETEROGENOUS SAMPLES IN FORENSIC APPLICATIONS. Forensic science international. Genetics Inci, F. n., Karaaslan, M. G., Gupta, R. n., Avadhani, A. n., Ogut, M. G., Atila, E. E., Duncan, G. n., Klevan, L. n., Demirci, U. n. 2020; 52: 102451

  Abstract

  Rapid and efficient processing of sexual assault evidence will accelerate forensic investigation and decrease casework backlogs. The standardized protocols currently used in forensic laboratories require the continued innovation to handle the increasing number and complexity of samples being submitted to forensic labs. Here, we present a new technique leveraging the integration of a bio-inspired oligosaccharide (i.e., Sialyl-LewisX) with magnetic beads that provides a rapid, inexpensive, and easy-to-use strategy that can potentially be adapted with current differential extraction practice in forensics labs. This platform (i) selectively captures sperm; (ii) is sensitive within the forensic cut-off; (iii) provides a cost effective solution that can be automated with existing laboratory platforms; and (iv) handles small volumes of sample (∼200 μL). This strategy can rapidly isolate sperm within 25 minutes of total processing that will prepare the extracted sample for downstream forensic analysis and ultimately help accelerate forensic investigation and reduce casework backlogs.

  View details for DOI 10.1016/j.fsigen.2020.102451

  View details for PubMedID 33556896

• Facilitating islet transplantation using a three-step approach with mesenchymal stem cells, encapsulation, and pulsed focused ultrasound. Stem cell research & therapy Razavi, M. n., Ren, T. n., Zheng, F. n., Telichko, A. n., Wang, J. n., Dahl, J. J., Demirci, U. n., Thakor, A. S. 2020; 11 (1): 405

  Abstract

  The aim of this study was to examine the effect of a three-step approach that utilizes the application of adipose tissue-derived mesenchymal stem cells (AD-MSCs), encapsulation, and pulsed focused ultrasound (pFUS) to help the engraftment and function of transplanted islets.In step 1, islets were co-cultured with AD-MSCs to form a coating of AD-MSCs on islets: here, AD-MSCs had a cytoprotective effect on islets; in step 2, islets coated with AD-MSCs were conformally encapsulated in a thin layer of alginate using a co-axial air-flow method: here, the capsule enabled AD-MSCs to be in close proximity to islets; in step 3, encapsulated islets coated with AD-MSCs were treated with pFUS: here, pFUS enhanced the secretion of insulin from islets as well as stimulated the cytoprotective effect of AD-MSCs.Our approach was shown to prevent islet death and preserve islet functionality in vitro. When 175 syngeneic encapsulated islets coated with AD-MSCs were transplanted beneath the kidney capsule of diabetic mice, and then followed every 3 days with pFUS treatment until day 12 post-transplantation, we saw a significant improvement in islet function with diabetic animals re-establishing glycemic control over the course of our study (i.e., 30 days). In addition, our approach was able to enhance islet engraftment by facilitating their revascularization and reducing inflammation.This study demonstrates that our clinically translatable three-step approach is able to improve the function and viability of transplanted islets.

  View details for DOI 10.1186/s13287-020-01897-z

  View details for PubMedID 32948247

• Medical Micro/Nanorobots in Precision Medicine Medical Micro/Nanorobots in Precision Medicine Soto, F., Wang, J., Ahmed, R., Demirci, U. 2020: 2002203

  View details for DOI 10.1002/advs.202002203

• Emerging organoid models: leaping forward in cancer research. Journal of hematology & oncology Fan, H., Demirci, U., Chen, P. 2019; 12 (1): 142

  Abstract

  Cancer heterogeneity is regarded as the main reason for the failure of conventional cancer therapy. The ability to reconstruct intra- and interpatient heterogeneity in cancer models is crucial for understanding cancer biology as well as for developing personalized anti-cancer therapy. Cancer organoids represent an emerging approach for creating patient-derived in vitro cancer models that closely recapitulate the pathophysiological features of natural tumorigenesis and metastasis. Meanwhile, cancer organoids have recently been utilized in the discovery of personalized anti-cancer therapy and prognostic biomarkers. Further, the synergistic combination of cancer organoids with organ-on-a-chip and 3D bioprinting presents a new avenue in the development of more sophisticated and optimized model systems to recapitulate complex cancer-stroma or multiorgan metastasis. Here, we summarize the recent advances in cancer organoids from a perspective of the in vitro emulation of natural cancer evolution and the applications in personalized cancer theranostics. We also discuss the challenges and trends in reconstructing more comprehensive cancer models for basic and clinical cancer research.

  View details for DOI 10.1186/s13045-019-0832-4

  View details for PubMedID 31884964

• Soft Ring-Shaped Cellu-Robots with Simultaneous Locomotion in Batches. Advanced materials (Deerfield Beach, Fla.) Ren, T., Chen, P., Gu, L., Ogut, M. G., Demirci, U. 2019: e1905713

  Abstract

  Untethered mini-robots can move single cells or aggregates to build complex constructs in confined spaces and may enable various biomedical applications such as regenerative repair in medicine and biosensing in bioengineering. However, a significant challenge is the ability to control multiple microrobots simultaneously in the same space to operate toward a common goal in a distributed operation. A locomotion strategy that can simultaneously guide the formation and operation of multiple robots in response to a common acoustic stimulus is developed. The scaffold-free cellu-robots comprise only highly packed cells and eliminate the influence of supportive materials, making them less cumbersome during locomotion. The ring shape of the cellu-robot contributes to anisotropic cellular interactions which induce radial cellular orientation. Under a single stimulus, several cellu-robots form predetermined complex structures such as bracelet-like ring-chains which transform into a single new living entity through cell-cell interactions, migration or cellular extensions between cellu-robots.

  View details for DOI 10.1002/adma.201905713

  View details for PubMedID 31773837

• Activate capture and digital counting (AC + DC) assay for protein biomarker detection integrated with a self-powered microfluidic cartridge. Lab on a chip Che, C., Li, N., Long, K. D., Aguirre, M. A., Canady, T. D., Huang, Q., Demirci, U., Cunningham, B. T. 2019

  Abstract

  We demonstrate a rapid, 2-step, and ultrasensitive assay approach for quantification of target protein molecules from a single droplet test sample. The assay is comprised of antibody-conjugated gold nanoparticles (AuNPs) that are "activated" when they are mixed with the test sample and bind their targets. The resulting liquid is passed through a microfluidic channel with a photonic crystal (PC) biosensor that is functionalized with secondary antibodies to the target biomarker, so that only activated AuNPs are captured. Utilizing recently demonstrated hybrid optical coupling between the plasmon resonance of the AuNP and the resonance of the PC, each captured AuNP efficiently quenches the resonant reflection of the PC, thus enabling the captured AuNPs to be digitally counted with high signal-to-noise. To achieve a 2-step assay process that is performed on a single droplet test sample without washing steps or active pump elements, controlled single-pass flow rate is obtained with an absorbing paper pad waste reservoir embedded in a microfluidic cartridge. We use the activate capture and digital counting (AC + DC) approach to demonstrate HIV-1 capsid antigen p24 detection from a 40 muL spiked-in human serum sample at a one thousand-fold dynamic range (1-103 pg mL-1) with only a 35-minute process that is compatible with point-of-care (POC) analysis. The AC + DC approach allows for ultrasensitive and ultrafast biomolecule detection, with potential applications in infectious disease diagnostics and early stage disease monitoring.

  View details for DOI 10.1039/c9lc00728h

  View details for PubMedID 31641717

• Utility of High-Sensitivity and Conventional Troponin in Patients Undergoing Transcatheter Aortic Valve Replacement: Incremental Prognostic Value to B-type Natriuretic Peptide. Scientific reports Kobayashi, Y., Kim, J. B., Moneghetti, K. J., Fischbein, M., Lee, A., Watkins, C. A., Yeung, A. C., Liang, D., Ozen, M. O., Demirci, U., Bowen, R., Fearon, W. F., Haddad, F. 2019; 9 (1): 14936

  Abstract

  High-sensitivity Troponin (hs-Tn) has emerged as a useful marker for patients with myocardial injury or heart failure. However, few studies have compared intermediate and hs-Tn in patients undergoing transcatheter aortic valve replacement (TAVR). Moreover, there remains uncertainty of which thresholds are the most useful for discriminating ventricular dysfunction or outcome. In this study we prospectively enrolled 105 patients with severe aortic stenosis (AS) who underwent TAVR as well as blood sampling for high-sensitivity (hs-TnI) and conventional troponin I (EXL-LOCI and RXL) assessment. Patients underwent comprehensive pre-procedure echocardiography. Ventricular dysfunction was defined using left ventricular mass index (LVMI), LV global longitudinal strain (LVGLS) and LV end-diastolic pressure. The mean age was 84.0±8.7 years old and 60% were male sex with mean transaortic pressure gradient of 50.1±16.0mmHg and AVA of 0.63±0.19cm2. When using a threshold of 6ng/L, 77% had positive hs-TnI while 27% had positive hs-TnI using recommended thresholds (16ng/L for female and 34ng/L for male). Troponin levels were higher in the presence of abnormal LV phenotypes. The strongest correlate of troponin was LVMI. During median follow-up of 375 days, 21 patients (20%) died. Lower threshold of hs-TnI and EXL-TnI was more discriminatory for overall mortality (Log-rank P=0.03 for both), while higher threshold of hs-TnI (p=0.75) and RXL-TnI were not (p=0.30). Combining hs-TnI and BNP improved to predict long-term outcome (p=0.004). In conclusion, hs-TnI levels correlated with the degree of LV dysfunction phenotypes. Furthermore, applying a lower threshold for hs-TnI performed better for outcome prediction than a recommended threshold in patients undergoing TAVR. Combining hs-TnI with BNP helped better risk stratification.

  View details for DOI 10.1038/s41598-019-51371-x

  View details for PubMedID 31624275

• Multi-stimuli-responsive programmable biomimetic actuator. Nature communications Dong, Y., Wang, J., Guo, X., Yang, S., Ozen, M. O., Chen, P., Liu, X., Du, W., Xiao, F., Demirci, U., Liu, B. 2019; 10 (1): 4087

  Abstract

  Untethered small actuators have various applications in multiple fields. However, existing small-scale actuators are very limited in their intractability with their surroundings, respond to only a single type of stimulus and are unable to achieve programmable structural changes under different stimuli. Here, we present a multiresponsive patternable actuator that can respond to humidity, temperature and light, via programmable structural changes. This capability is uniquely achieved by a fast and facile method that was used to fabricate a smart actuator with precise patterning on a graphene oxide film by hydrogel microstamping. The programmable actuator can mimic the claw of a hawk to grab a block, crawl like an inchworm, and twine around and grab the rachis of a flower based on their geometry. Similar to the large- and small-scale robots that are used to study locomotion mechanics, these small-scale actuators can be employed to study movement and biological and living organisms.

  View details for DOI 10.1038/s41467-019-12044-5

  View details for PubMedID 31501430

• Plasmonic-based platforms for diagnosis of infectious diseases at the point-of-care. Biotechnology advances Li, Z., Leustean, L., Inci, F., Zheng, M., Demirci, U., Wang, S. 2019: 107440

  Abstract

  Infectious diseases such as HIV-1, TB, HBV and malaria still exert a tremendous health burden on the developing world, requiring rapid, simple and inexpensive diagnostics for on-site diagnosis and treatment monitoring. However, traditional diagnostic methods such as nucleic acid tests (NATs) and enzyme linked immunosorbent assays (ELISA) cannot be readily implemented in point-of-care (POC) settings. Recently, plasmonic-based biosensors have emerged, offering an attractive solution to manage infectious diseases in the developing world since they can achieve rapid, real-time and label-free detection of various pathogenic biomarkers. Via the principle of plasmonic-based optical detection, a variety of biosensing technologies such as surface plasmon resonance (SPR), localized surface plasmon resonance (LSPR), colorimetric plasmonic assays, and surface enhanced Raman spectroscopy (SERS) have emerged for early diagnosis of HIV-1, TB, HBV and malaria. Similarly, plasmonic-based colorimetric assays have also been developed, with the capability of multiplexing and cellphone integration, which is well suited for POC testing in the developing world. Herein, we present a comprehensive review on recent advances in surface chemistry, substrate fabrication and microfluidic integration for the development of plasmonic-based biosensors, aiming at rapid management of infectious diseases at the POC, and thus, improving global health.

  View details for DOI 10.1016/j.biotechadv.2019.107440

  View details for PubMedID 31476421

• Amplification of nuclear deformation of breast cancer cells by seeding on micropatterned surfaces to better distinguish their malignancies. Colloids and surfaces. B, Biointerfaces Antmen, E., Demirci, U., Hasirci, V. 2019; 183: 110402

  Abstract

  Information about the mechanical properties of cancer cells leads to new insights about their malignancy levels. The more flexible the cancer cells and their nuclei are, the more aggressive and invasive they are. Flexibility is a result of composition and properties of molecular constituents of cells and its extent is expressed by deformation. Differences in the mechanical properties could be modulated by topography and chemistry of the substrate. In this study, the main hypothesis is that the difference in the mechanical properties of malignant and benign breast cancer cells could be used as a discriminator of these cells and reflected by the extent of nuclear deformation on micropatterned substrates. We compared benign (MCF10A), malignantnoninvasive (MCF7), and malignant highly invasive (MDAMB231) breast cancer cell lines using their nuclear deformability on micropatterned surfaces designed with square prism-shaped micropillars of poly(methyl methacrylate) (PMMA) (8 mum high, 4 * 4 mum2 area, 4 mum gap). Several shape descriptors (circularity, solidity, roundness, aspect ratio) were used to analyze nuclear deformation. We were able to discriminate the three cells when the descriptor circularity and hydrophobic micropatterned surfaces were used. The cells showed nuclear deformability in the order following the extent of their malignancies. The most aggressive cell, MDAMB231, had the lowest circularity value, 0.37, whereas the noninvasive malignant, MCF7, and benign, MCF10A, cells had higher values 0.47 and 0.77, respectively. Mechanism of the deformation was shown at the molecular level that the expression of Lamin A/C and Nesprin-2 genes decreased with increased nuclear deformation. In summary, biomechanical properties of cells can provide useful information about their cancer state and they can be reflected in the biological markers.

  View details for DOI 10.1016/j.colsurfb.2019.110402

  View details for PubMedID 31398621

• Square prism micropillars on poly(methyl methacrylate) surfaces modulate the morphology and differentiation of human dental pulp mesenchymal stem cells COLLOIDS AND SURFACES B-BIOINTERFACES Hasturk, O., Ermis, M., Demirci, U., Hasirci, N., Hasirci, V. 2019; 178: 44–55

  View details for DOI 10.1016/j.colsurfb.2019.02.037

  View details for Web of Science ID 000471084400006

• Density Based Characterization of Mechanical Cues on Cancer Cells Using Magnetic Levitation ADVANCED HEALTHCARE MATERIALS Baday, M., Ercal, O., Sahan, A., Sahan, A., Ercal, B., Inan, H., Demirci, U. 2019; 8 (10)

  View details for DOI 10.1002/adhm.201801517

  View details for Web of Science ID 000468803300015

• Microfluidic Chip for Detection of Fungal Infections. ACS omega Asghar, W., Sher, M., Khan, N. S., Vyas, J. M., Demirci, U. 2019; 4 (4): 7474–81

  Abstract

  Fungal infections can lead to severe clinical outcomes such as multiple organ failure and septic shock. Rapid detection of fungal infections allows clinicians to treat patients in a timely manner and improves clinical outcomes. Conventional detection methods include blood culture followed by plate culture and polymerase chain reaction. These methods are time-consuming and require expensive equipment, hence, they are not suitable for point-of-care and clinical settings. There is an unmet need to develop a rapid and inexpensive detection method for fungal infections such as candidemia. We developed an innovative immuno-based microfluidic device that can rapidly detect and capture Candida albicans from phosphate-buffered saline (PBS) and human whole blood. Our microchip technology showed an efficient capture of C. albicans in PBS with an efficiency of 61-78% at various concentrations ranging from 10 to 105 colony-forming units per milliliter (cfu/mL). The presented microfluidic technology will be useful to screen for various pathogens at the point-of-care and clinical settings.

  View details for PubMedID 31080939

• Density Based Characterization of Mechanical Cues on Cancer Cells Using Magnetic Levitation. Advanced healthcare materials Baday, M., Ercal, O., Sahan, A. Z., Sahan, A., Ercal, B., Inan, H., Demirci, U. 2019: e1801517

  Abstract

  Extracellular matrix (ECM) stiffness is correlated to malignancy and invasiveness of cancer cells. Although the mechanism of change is unclear, mechanical signals from the ECM may affect physical properties of cells such as their density profile. The current methods, such as Percoll density-gradient centrifugation, are unable to detect minute density differences. A magnetic levitation device is developed (i.e., MagDense platform) where cells are levitated in a magnetic gradient allowing them to equilibrate to a levitation height that corresponds to their unique cellular density. In application of this system, MDA-MB-231 breast and A549 lung cancer cells are cultured and overall differences in cell density are observed in response to increasing collagen fiber density. Overall, density values are significantly more spread out for MDA-MB-231 cells extracted from the 1.44 mg mL-1 collagen gels compared to those from 0.72 mg mL-1 collagen, whereas no significant difference with A549 cell lines is observed. The MagDense platform can determine differences in cellular densities under various microenvironmental conditions. The imaging of cancer cells in a magnetic levitation device serves as a unique tool to observe changes in phenotypic properties of cancer cells as they relate to micromechanical cues encoded by the ECM.

  View details for PubMedID 30946539

• Loss of Endothelium-Derived Wnt5a Is Associated With Reduced Pericyte Recruitment and Small Vessel Loss in Pulmonary Arterial Hypertension CIRCULATION Yuan, K., Shamskhou, E. A., Orcholski, M. E., Nathan, A., Reddy, S., Honda, H., Mani, V., Zeng, Y., Ozen, M. O., Wang, L., Demirci, U., Tian, W., Nicolls, M. R., Perez, V. 2019; 139 (14): 1710–24

  View details for DOI 10.1161/CIRCULATIONAHA.118.037642

  View details for Web of Science ID 000469320000018

• Microfluidic Chip for Detection of Fungal Infections ACS OMEGA Asghar, W., Sher, M., Khan, N. S., Vyas, J. M., Demirci, U. 2019; 4 (4): 7474–81

  View details for DOI 10.1021/acsomega.9b00499

  View details for Web of Science ID 000466552500150

• Bioinspired Preservation of Natural Killer Cells for Cancer Immunotherapy. Advanced science (Weinheim, Baden-Wurttemberg, Germany) El Assal, R., Abou-Elkacem, L., Tocchio, A., Pasley, S., Matosevic, S., Kaplan, D. L., Zylberberg, C., Demirci, U. 2019; 6 (6): 1802045

  Abstract

  The ability to cryopreserve natural killer (NK) cells has a significant potential in modern cancer immunotherapy. Current cryopreservation protocols cause deterioration in NK cell viability and functionality. This work reports the preservation of human cytokine-activated NK cell viability and function following cryopreservation using a cocktail of biocompatible bioinspired cryoprotectants (i.e., dextran and carboxylated ε-poly-L-lysine). Results demonstrate that the recovered NK cells after cryopreservation and rewarming maintain their viability immediately after thawing at a comparable level to control (dimethyl sulfoxide-based cryopreservation). Although, their viability drops in the first day in culture compared to controls, the cells grow back to a comparable level to controls after 1 week in culture. In addition, the anti-tumor functional activity of recovered NK cells demonstrates higher cytotoxic potency against leukemia cells compared to control. This approach presents a new direction for NK cell preservation, focusing on function and potentially enabling storage and distribution for cancer immunotherapy.

  View details for DOI 10.1002/advs.201802045

  View details for PubMedID 30937270

  View details for PubMedCentralID PMC6425501

• MICROVESICLES LARGER THAN 200NM RESCUE CARDIOMYOCYTES FROM DOXORUBICIN INJURY IN A PATIENT-SPECIFIC MODEL OF ANTHRACYCLINE INDUCED CARDIOMYOPATHY O'Brien, C., Shi, L., Ozgun, M., Vaskova, E., Santoso, M., Jung, J., Ikeda, G., Demirci, U., Yang, P. ELSEVIER SCIENCE INC. 2019: 688

  View details for Web of Science ID 000460565900688

• INDEPENDENT PROGNOSTIC VALUES OF CLINICAL RISK SCORES, RIGHT VENTRICULAR SYSTOLIC PRESSURE, AND N-TERMINAL PRO-B-TYPE PEPTIDE IN HEART FAILURE WITH PRESERVED EJECTION FRACTION: INSIGHTS FROM SUPERVISED AND UNSUPERVISED MODELS Tremblay-Gravel, M., Kobayashi, Y., Boralkar, K., Li, X., Bouajila, S., Nishi, T., Amsallem, M., Moneghetti, K., Selej, M., Ozen, M., Demirci, U., Ashley, E. A., Wheeler, M., Knowlton, K., Kouznetsova, T., Haddad, F. ELSEVIER SCIENCE INC. 2019: 718

  View details for Web of Science ID 000460565900718

• Square prism micropillars on poly(methyl methacrylate) surfaces modulate the morphology and differentiation of human dental pulp mesenchymal stem cells. Colloids and surfaces. B, Biointerfaces Hasturk, O., Ermis, M., Demirci, U., Hasirci, N., Hasirci, V. 2019; 178: 44–55

  Abstract

  Use of soluble factors is the most common strategy to induce osteogenic differentiation of mesenchymal stem cells (MSCs) in vitro, but it may raise potential side effects in vivo. The topographies of the substrate surfaces affect cell behavior, and this could be a promising approach to guide stem cell differentiation. Micropillars have been reported to modulate cellular and subcellular shape, and it is particularly interesting to investigate whether these changes in cell morphology can modulate gene expression and lineage commitment without chemical induction. In this study, poly(methyl methacrylate) (PMMA) films were decorated with square prism micropillars with different lateral dimensions (4, 8 and 16mum), and the surface wettability of the substrates was altered by oxygen plasma treatment. Both, pattern dimensions and hydrophilicity, were found to affect the attachment, proliferation, and most importantly, gene expression of human dental pulp mesenchymal stem cells (DPSCs). Decreasing the pillar width and interpillar spacing of the square prism pillars enhanced cell attachment, cell elongation, and deformation of nuclei, but reduced early proliferation rate. Surfaces with 4 or 8mum wide pillars/gaps upregulated the expression of early bone-marker genes and mineralization over 28 days of culture. Exposure to oxygen plasma increased wettability and promoted cell attachment and proliferation but delayed osteogenesis. Our findings showed that surface topography and chemistry are very useful tools in controlling cell behavior on substrates and they can also help create better implants. The most important finding is that hydrophobic micropillars on polymeric substrate surfaces can be exploited in inducing osteogenic differentiation of MSCs without any differentiation supplements.

  View details for PubMedID 30826553

• Engineered natural and synthetic polymer surfaces induce nuclear deformation in osteosarcoma cells JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART B-APPLIED BIOMATERIALS Antmen, E., Ermis, M., Demirci, U., Hasirci, V. 2019; 107 (2): 366–76

  Abstract

  Cell-substrate interactions involve constant probing of microenvironment by cells. One of the responses of cells to environmental cues is to change the conformation of their cytoplasm and nucleus. We hypothesized that surface chemistry and topography could be engineered to make these differences significant enough. When designing the substrates that would accentuate these differences, we prepared surfaces carrying cell adhesive biological cues arranged in specific patterns. Collagen type I and poly(lactic acid-co-glycolic acid) (PLGA) were used to represent substrates with biological cues and those without, and these materials were decorated with four square prism micropillars with different dimensions. The nuclear deformations were analyzed using some descriptors. Nucleus area and solidity were the best descriptors in distinguishing the substrates in terms of biological cues, while nucleus area, solidity, and circularity were more sensitive to the interpillar distances. Another distinguishing factor tested was the duration of contact. Nucleus area was the only descriptor sensitive to nuclear deformation change with time. PLGA was more suitable in nuclear conformation analysis while collagen was better in cell adhesion and proliferation. These deformations lead to changes in the molecular processes and further studies are needed to better understand cell mechanobiology. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 107B: 366-376, 2019.

  View details for PubMedID 29663651

• 3-D geometry and irregular connectivity dictate neuronal firing in frequency domain and synchronization. Biomaterials Ren, T., Grosshauser, B., Sridhar, K., Nieland, T. J., Tocchio, A., Schepers, U., Demirci, U. 2019; 197: 171–81

  Abstract

  The replication of the complex structure and three dimensional (3-D) interconnectivity of neurons in the brain is a great challenge. A few 3-D neuronal patterning approaches have been developed to mimic the cell distribution in the brain but none have demonstrated the relationship between 3-D neuron patterning and network connectivity. Here, we used photolithographic crosslinking to fabricate in vitro 3-D neuronal structures with distinct sizes, shapes or interconnectivities, i.e., milli-blocks, micro-stripes, separated micro-blocks and connected micro-blocks, which have spatial confinement from "Z" dimension to "XYZ" dimension. During a 4-week culture period, the 3-D neuronal system has shown high cell viability, axonal, dendritic, synaptic growth and neural network activity of cortical neurons. We further studied the calcium oscillation of neurons in different 3-D patterns and used signal processing both in Fast Fourier Transform (FFT) and time domain (TD) to model the fluorescent signal variation. We observed that the firing frequency decreased as the spatial confinement in 3-D system increased. Besides, the neuronal synchronization significantly decreased by irregularly connecting micro-blocks, indicating that network connectivity can be adjusted by changing the linking conditions of 3-D gels. Earlier works showed the importance of 3-D culture over 2-D in terms of cell growth. Here, we showed that not only 3-D geometry over 2-D culture matters, but also the spatial organization of cells in 3-D dictates the neuronal firing frequency and synchronicity.

  View details for DOI 10.1016/j.biomaterials.2019.01.017

  View details for PubMedID 30660993

• Epithelial-to-Mesenchymal Transition (EMT) and Drug Response in Dynamic Bioengineered Lung Cancer Microenvironment ADVANCED BIOSYSTEMS Mani, V., Lyu, Z., Kumar, V., Ercal, B., Chen, H., Malhotra, S., Demirci, U. 2019; 3 (1)

  View details for DOI 10.1002/adbi.201800223

  View details for Web of Science ID 000455812800003

• Epithelial-to-Mesenchymal Transition (EMT) and Drug Response in Dynamic Bioengineered Lung Cancer Microenvironment. Advanced biosystems Mani, V., Lyu, Z., Kumar, V., Ercal, B., Chen, H., Malhotra, S. V., Demirci, U. 2019; 3 (1): e1800223

  Abstract

  Tumor microenvironment and the interplay of physical and mechanical forces are key determinants of cancer initiation, progression, and response to drug treatment. However, the impact of tumor microenvironment on cancer progression is poorly understood, in large due to the lack of in vitro models that recapitulate the physical aspects of tumor microenvironment. Herein, a simple, dynamic 3D nonsmall cell lung carcinoma culture using a multichannel microfluidic model platform is developed for evaluating the contribution of flow-induced hydrodynamic shear stress on epithelial-to-mesenchymal transition (EMT). It is found that flow induces changes in cellular morphology and EMT in 2D and 3D when lung cancer A549 cells are cultured on a microfluidic chip under laminar flow for 4-5 days compared to traditional static cultures. The role of dynamic cell culture on chemotherapeutic effects is monitored. Drug response with an existing anti-cancer drug, e.g., erlotinib and an investigational drug (NSC-750212), shows distinct cytotoxic effects in flow compared to static cultures, suggesting a potential influence of flow on drug efficacy in 2D and 3D models. The platform demonstrates the ability to create a dynamic microscale tumor model, which could be explored as a tool for early drug screening and treatment monitoring in cancer and other diseases.

  View details for DOI 10.1002/adbi.201800223

  View details for PubMedID 32627339

• Real-time Biosensing of Proteins on a DVD Nanoplasmonic Grating Ahmed, R., Ozen, M., Inci, F., Karaaslan, M., Henrich, T. J., Demirci, U., VoDinh, T., Ho, H. P., Ray, K. SPIE-INT SOC OPTICAL ENGINEERING. 2019

  View details for DOI 10.1117/12.2508624

  View details for Web of Science ID 000482226200010

• Bioinspired Preservation of Natural Killer Cells for Cancer Immunotherapy Advanced Science El Assal, R., Abou-Elkacem, L., Tocchio, A., Pasley, S., Matosevic, S., Kaplan, D., Zylberberg, C., Demirci, U. 2019

  View details for DOI 10.1002/advs.201802045

• MICROSCALE MAGNETIC LEVITATION FOR MULTIPLEXED ANALYSIS OF MALARIA-INFECTED BLOOD SAMPLES IN RESOURCE-LIMITED SETTINGS Deshmukh, S. S., Durmus, N., Greenhouse, B., Egan, E., Demirci, U. AMER SOC TROP MED & HYGIENE. 2019: 130–31

  View details for Web of Science ID 000507364502427

• Approaching Higher Dimension Imaging Data Using Cluster-Based Hierarchical Modeling in Patients with Heart Failure Preserved Ejection Fraction. Scientific reports Kobayashi, Y. n., Tremblay-Gravel, M. n., Boralkar, K. A., Li, X. n., Nishi, T. n., Amsallem, M. n., Moneghetti, K. J., Bouajila, S. n., Selej, M. n., Ozen, M. O., Demirci, U. n., Ashley, E. n., Wheeler, M. n., Knowlton, K. U., Kouznetsova, T. n., Haddad, F. n. 2019; 9 (1): 10431

  Abstract

  Heart failure with preserved ejection fraction (HFpEF) is a major cause of morbidity and mortality, accounting for the majority of heart failure (HF) hospitalization. To identify the most complementary predictors of mortality among clinical, laboratory and echocardiographic data, we used cluster based hierarchical modeling. Using Stanford Translational Research Database, we identified patients hospitalized with HFpEF between 2005 and 2016 in whom echocardiogram and NT-proBNP were both available at the time of admission. Comprehensive echocardiographic assessment including left ventricular longitudinal strain (LVLS), right ventricular function and right ventricular systolic pressure (RVSP) was performed. The outcome was defined as all-cause mortality. Among patients identified, 186 patients with complete echocardiographic assessment were included in the analysis. The cohort included 58% female, with a mean age of 78.7 ± 13.5 years, LVLS of -13.3 ± 2.5%, an estimated RVSP of 38 ± 13 mmHg. Unsupervised cluster analyses identified six clusters including ventricular systolic-function cluster, diastolic-hemodynamic cluster, end-organ function cluster, vital-sign cluster, complete blood count and sodium clusters. Using a stepwise hierarchical selection from each cluster, we identified NT-proBNP (standard hazard ratio [95%CI] = 1.56 [1.17-2.08]) and RVSP (1.37 [1.09-1.78]) as independent correlates of outcome. When adding these parameters to the well validated Get with the Guideline Heart Failure risk score, the Chi-square was significantly improved (p = 0.01). In conclusion, NT-proBNP and RVSP were independently predictive in HFpEF among clinical, imaging, and biomarker parameters. Cluster-based hierarchical modeling may help identify the complementally predictive parameters in small cohorts with higher dimensional clinical data.

  View details for DOI 10.1038/s41598-019-46873-7

  View details for PubMedID 31320698

• Biochemical Gradients to Generate 3D Heterotypic-Like Tissues with Isotropic and Anisotropic Architectures ADVANCED FUNCTIONAL MATERIALS Canadas, R. F., Ren, T., Marques, A. P., Oliveira, J. M., Reis, R. L., Demirci, U. 2018; 28 (48)

  View details for DOI 10.1002/adfm.201804148

  View details for Web of Science ID 000451118800017

• A Novel On-Chip Method for Differential Extraction of Sperm in Forensic Cases ADVANCED SCIENCE Inci, F., Ozen, M. O., Saylan, Y., Miansari, M., Cimen, D., Dhara, R., Chinnasamy, T., Yuksekkaya, M., Filippini, C., Kumar, D., Calamak, S., Yesil, Y., Durmus, N., Duncan, G., Klevan, L., Demirci, U. 2018; 5 (9): 1800121

  Abstract

  One out of every six American women has been the victim of a sexual assault in their lifetime. However, the DNA casework backlog continues to increase outpacing the nation's capacity since DNA evidence processing in sexual assault casework remains a bottleneck due to laborious and time-consuming differential extraction of victim's and perpetrator's cells. Additionally, a significant amount (60-90%) of male DNA evidence may be lost with existing procedures. Here, a microfluidic method is developed that selectively captures sperm using a unique oligosaccharide sequence (Sialyl-LewisX), a major carbohydrate ligand for sperm-egg binding. This method is validated with forensic mock samples dating back to 2003, resulting in 70-92% sperm capture efficiency and a 60-92% reduction in epithelial fraction. Captured sperm are then lysed on-chip and sperm DNA is isolated. This method reduces assay-time from 8 h to 80 min, providing an inexpensive alternative to current differential extraction techniques, accelerating identification of suspects and advancing public safety.

  View details for PubMedID 30250782

• Microfluidic sorting selects sperm for clinical use with reduced DNA damage compared to density gradient centrifugation with swim-up in split semen samples HUMAN REPRODUCTION Quinn, M. M., Jalalian, L., Ribeiro, S., Ona, K., Demirci, U., Cedars, M. I., Rosen, M. P. 2018; 33 (8): 1388–93

  Abstract

  Does microfluidic sorting improve the selection of sperm with lower DNA fragmentation over standard density-gradient centrifugation?Microfluidic sorting of unprocessed semen allows for the selection of clinically usable, highly motile sperm with nearly undetectable levels of DNA fragmentation.Microfluidic devices have been explored to sort motile and morphologically normal sperm from a raw sample without centrifugation; however, it is uncertain whether DNA damage is reduced in this process.This is a blinded, controlled laboratory study of differences in standard semen analysis parameters and the DNA fragmentation index (DFI) in split samples from infertile men (n = 70) that were discarded after routine semen analysis at an academic medical center.Sperm concentration, progressive motility and forward progression were assessed by microscopic examination. For each sample, the unprocessed semen was tested for DNA fragmentation and split for processing by density-gradient centrifugation with swim-up or sorting by a microfluidic chip. DNA fragmentation was assessed in unprocessed and processed samples by sperm chromatin dispersion assay. The DFI was calculated, from up to 300 cells per slide, as the number of cells with fragmented DNA divided by the number of cells counted per slide.The median DFI in unprocessed samples was 21% (interquartile range (IQR): 14-30). In paired analyses of all samples, those processed by the microfluidic chip demonstrated significantly decreased DFI compared to those processed by density-gradient centrifugation (P = 0.0029) and unprocessed samples (P < 0.0001). The median DFI for chip specimens was 0% (IQR: 0-2.4) while those processed by density-gradient centrifugation had a median DFI of 6% (IQR: 2-11). Unprocessed samples in the highest DFI quartile (DFI range: 31-40%) had a median DFI of 15% (IQR: 11-19%) after density-gradient centrifugation and DFI of 0% (IQR: 0-1.9%) after processing with the microfluidic chip (P = 0.02).While a high DFI has been associated with poor outcomes with IVF/ICSI, there are limited data illustrating improvements in clinical outcomes with a reduction in DFI. As this study utilized discarded, non-clinical samples, clinical outcomes data are not available.While microfluidic sorting of unprocessed semen allowed for the selection of clinically usable, highly motile sperm with nearly undetectable levels of DNA fragmentation, standard processing by density-gradient centrifugation with swim-up did not increase DNA fragmentation in an infertile population. The proposed microfluidic technology offers a flow-free approach to sort sperm, requiring no peripheral equipment or filtration step, while minimizing hands-on time.No external funding to declare. Utkan Demirci, PhD is the Co-founder and Scientific Advisor for DxNow Inc., LevitasBio Inc. and Koek Biotech. Mitchell Rosen, MD is a member of the Clinical Advisory Board for DxNow Inc.

  View details for PubMedID 30007319

• Tunable anisotropic networks for 3-D oriented neural tissue models. Biomaterials Canadas, R. F., Ren, T., Tocchio, A., Marques, A. P., Oliveira, J. M., Reis, R. L., Demirci, U. 2018; 181: 402–14

  Abstract

  Organized networks are common in nature showing specific tissue micro-architecture, where cells can be found isotropically or anisotropically distributed in characteristic arrangements and tissue stiffness. However, when addressing an in vitro tissue model, it is challenging to grant control over mechanical properties while achieving anisotropic porosity of polymeric networks, especially in three-dimensional systems (3-D). While progress was achieved organizing cells in two-dimension (2-D), fabrication methods for aligned networks in 3-D are limited. Here, we describe the use of a biomimetic extra-cellular matrix system allowing programming of anisotropic structures into precisely advancing pore diameters in 3-D. Using control over polymeric composition, crosslinking directionality and freezing gradient dynamics, we revealed a mechanism to top-down biofabricate 3-D structures with tunable micro-porosity capable of directing cellular responses at millimeter scale such as axonal anisotropic outgrowth that is a unique characteristic of the brain cortex. Further, we showed the unique integration of this method with a microfluidic system establishing a neural-endothelial heterotypic conjugation, which can potentially be broadly applied to multiple organ systems.

  View details for PubMedID 30138793

• Ultrastructural brain abnormalities and associated behavioral changes in mice after low-intensity blast exposure BEHAVIOURAL BRAIN RESEARCH Song, H., Konan, L. M., Cui, J., Johnson, C. E., Langenderfer, M., Grant, D., Ndam, T., Simonyi, A., White, T., Demirci, U., Mott, D. R., Schwer, D., Hubler, G. K., Cernak, I., DePalma, R. G., Gu, Z. 2018; 347: 148–57

  Abstract

  Explosive blast-induced mild traumatic brain injury (mTBI), a "signature wound" of recent military conflicts, commonly affects service members. While past blast injury studies have provided insights into TBI with moderate- to high-intensity explosions, the impact of primary low-intensity blast (LIB)-mediated pathobiology on neurological deficits requires further investigation. Our prior considerations of blast physics predicted ultrastructural injuries at nanoscale levels. Here, we provide quantitative data using a primary LIB injury murine model exposed to open field detonation of 350 g of high-energy explosive C4. We quantified ultrastructural and behavioral changes up to 30 days post blast injury (DPI). The use of an open-field experimental blast generated a primary blast wave with a peak overpressure of 6.76 PSI (46.6 kPa) at a 3-m distance from the center of the explosion, a positive phase duration of approximate 3.0 milliseconds (ms), a maximal impulse of 8.7 PSI × ms and a sharp rising time of 9 × 10-3 ms, with no apparent impact/acceleration in exposed animals. Neuropathologically, myelinated axonal damage was observed in blast-exposed groups at 7 DPI. Using transmission electron microscopy, we observed and quantified myelin sheath defects and mitochondrial abnormalities at 7 and 30 DPI. Inverse correlations between blast intensities and neurobehavioral outcomes including motor activities, anxiety levels, nesting behavior, spatial learning and memory occurred. These observations uncover unique ultrastructural brain abnormalities and associated behavioral changes due to primary blast injury and provide key insights into its pathogenesis and potential treatment.

  View details for PubMedID 29526786

• Scaffold-free, label-free and nozzle-free biofabrication technology using magnetic levitational assembly BIOFABRICATION Parfenov, V. A., Koudan, E. V., Bulanova, E. A., Karalkin, P. A., Pereira, F. S., Norkin, N. E., Knyazeva, A. D., Gryadunova, A. A., Petrov, O. F., Vasiliev, M. M., Myasnikov, M. I., Chernikov, V. P., Kasyanov, V. A., Marchenkov, A., Brakke, K., Khesuani, Y. D., Demirci, U., Mironov, V. A. 2018; 10 (3): 034104

  Abstract

  Tissue spheroids have been proposed as building blocks in 3D biofabrication. Conventional magnetic force-driven 2D patterning of tissue spheroids requires prior cell labeling by magnetic nanoparticles, meanwhile a label-free approach for 3D magnetic levitational assembly has been introduced. Here we present first time report on rapid assembly of 3D tissue construct using scaffold-free, nozzle-free and label-free magnetic levitation of tissue spheroids. Chondrospheres of standard size, shape and capable to fusion have been biofabricated from primary sheep chondrocytes using non-adhesive technology. Label-free magnetic levitation was performed using a prototype device equipped with permanent magnets in presence of gadolinium (Gd3+) in culture media, which enables magnetic levitation. Mathematical modeling and computer simulations were used for prediction of magnetic field and kinetics of tissue spheroids assembly into 3D tissue constructs. First, we used polystyrene beads to simulate the assembly of tissue spheroids and to determine the optimal settings for magnetic levitation in presence of Gd3+. Second, we proved the ability of chondrospheres to assemble rapidly into 3D tissue construct in the permanent magnetic field in the presence of Gd3+. Thus, scaffold- and label-free magnetic levitation of tissue spheroids is a promising approach for rapid 3D biofabrication and attractive alternative to label-based magnetic force-driven tissue engineering.

  View details for DOI 10.1088/1758-5090/aac900

  View details for Web of Science ID 000435633000001

  View details for PubMedID 29848793

• An Embryonic and Induced Pluripotent Stem Cell Model for Ovarian Granulosa Cell Development and Steroidogenesis REPRODUCTIVE SCIENCES Lipskind, S., Lindsey, J. S., Gerami-Naini, B., Eaton, J. L., O'Connell, D., Kiezun, A., Ho, J. K., Ng, N., Parasar, P., Ng, M., Nickerson, M., Demirci, U., Maas, R., Anchan, R. M. 2018; 25 (5): 712–26

  Abstract

  Embryoid bodies (EBs) can serve as a system for evaluating pluripotency, cellular differentiation, and tissue morphogenesis. In this study, we use EBs derived from mouse embryonic stem cells (mESCs) and human amniocyte-derived induced pluripotent stem cells (hAdiPSCs) as a model for ovarian granulosa cell (GC) development and steroidogenic cell commitment. We demonstrated that spontaneously differentiated murine EBs (mEBs) and human EBs (hEBs) displayed ovarian GC markers, such as aromatase (CYP19A1), FOXL2, AMHR2, FSHR, and GJA1. Comparative microarray analysis identified both shared and unique gene expression between mEBs and the maturing mouse ovary. Gene sets related to gonadogenesis, lipid metabolism, and ovarian development were significantly overrepresented in EBs. Of the 29 genes, 15 that were differentially regulated in steroidogenic mEBs displayed temporal expression changes between embryonic, postnatal, and mature ovarian tissues by polymerase chain reaction. Importantly, both mEBs and hEBs were capable of gonadotropin-responsive estradiol (E2) synthesis in vitro (217-759 pg/mL). Live fluorescence-activated cell sorting-sorted AMHR2+ granulosa-like cells from mEBs continued to produce E2 after purification (15.3 pg/mL) and secreted significantly more E2 than AMHR2- cells (8.6 pg/mL, P < .05). We conclude that spontaneously differentiated EBs of both mESC and hAdiPSC origin can serve as a biologically relevant model for ovarian GC differentiation and steroidogenic cell commitment. These cells should be further investigated for therapeutic uses, such as stem cell-based hormone replacement therapy and in vitro maturation of oocytes.

  View details for DOI 10.1177/1933719117725814

  View details for Web of Science ID 000435535900010

  View details for PubMedID 28854867

• Evaluation of an ovary-on-a-chip in large mammalian models: Species specificity and influence of follicle isolation status JOURNAL OF TISSUE ENGINEERING AND REGENERATIVE MEDICINE Nagashima, J. B., El Assal, R., Songsasen, N., Demirci, U. 2018; 12 (4): E1926–E1935

  Abstract

  The ability to grow oocytes from immature ovarian follicles in vitro has significant potential for fertility preservation; yet, it has proved challenging in large mammalian species due to the complex metabolic needs and long-term culture requirements. Currently, follicular incubations are based on a "static" system with manual exchange of medium. Despite the numerous advantages of conventional culturing approaches, recapitulating the native microenvironment and supporting the survival of ovarian follicles from large mammalian species still represent challenges. In this study, we utilized an innovative, dynamic microfluidic system to support the in vitro survival of domestic cat and dog follicles enclosed within the ovarian cortex or isolated from ovarian cortex. Results indicate both species-specific and tissue type-specific differences in response to microfluidic culture. Domestic cat but not dog ovarian cortical tissues maintained viability under flow similar to conventional agarose gel controls. Preantral stage isolated follicles from both species that grew most favourably in conventional alginate bead culture, but overall, there was no influence of culture system on expression of follicle development or oocyte health markers. This system represents an important exploration toward the development of an improved ovarian in vitro culture system of large mammalian species (e.g., cats and dogs), which has potential applications for fertility preservation, reproductive toxicology, and endangered mammal conservation efforts.

  View details for PubMedID 29222841

  View details for PubMedCentralID PMC5906142

• A decade of progress in liver regenerative medicine BIOMATERIALS Zhang, J., Zhao, X., Liang, L., Li, J., Demirci, U., Wang, S. 2018; 157: 161–76

  Abstract

  Liver diseases can be caused by viral infection, metabolic disorder, alcohol consumption, carcinoma or injury, chronically progressing to end-stage liver disease or rapidly resulting in acute liver failure. In either situation, liver transplantation is most often sought for life saving, which is, however, significantly limited by severe shortage of organ donors. Until now, tremendous multi-disciplinary efforts have been dedicated to liver regenerative medicine, aiming at providing transplantable cells, microtissues, or bioengineered whole liver via tissue engineering, or maintaining partial liver functions via extracorporeal support. In both directions, new compatible biomaterials, stem cell sources, and bioengineering approaches have fast-forwarded liver regenerative medicine towards potential clinical applications. Another important progress in this field is the development of liver-on-a-chip technologies, which enable tissue engineering, disease modeling, and drug testing under biomimetic extracellular conditions. In this review, we aim to highlight the last decade's progress in liver regenerative medicine from liver tissue engineering, bioartificial liver devices (BAL), to liver-on-a-chip platforms, and then to present challenges ahead for further advancement.

  View details for PubMedID 29274550

• A MAGNETIC LEVITATION PLATFORM FOR THE ISOLATION OF MATURE SPERM FROM TESE/TESA SAMPLES Durmus, G., Gupta, R., Badamjav, O., Reddy, V., Eisenberg, M. L., Behr, B., Demirci, U. ELSEVIER SCIENCE INC. 2018: E26–E27

  View details for Web of Science ID 000427891800035

• An Automated Microfluidic Assay for Photonic Crystal Enhanced Detection and Analysis of an Antiviral Antibody Cancer Biomarker in Serum. IEEE sensors journal Race, C. M., Kwon, L. E., Foreman, M. T., Huang, Q., Inan, H., Kesiraju, S., Le, P., Lim, S. J., Smith, A. M., Zangar, R. C., Demirci, U., Anderson, K. S., Cunningham, B. T. 2018; 18 (4): 1464-1473

  Abstract

  We report on the implementation of an automated platform for detecting the presence of an antibody biomarker for human papillomavirus-associated oropharyngeal cancer from a single droplet of serum, in which a nanostructured photonic crystal surface is used to amplify the output of a fluorescence-linked immunosorbent assay. The platform is comprised of a microfluidic cartridge with integrated photonic crystal chips that interfaces with an assay instrument that automates the introduction of reagents, wash steps, and surface drying. Upon assay completion, the cartridge interfaces with a custom laser-scanning instrument that couples light into the photonic crystal at the optimal resonance condition for fluorescence enhancement. The instrument is used to measure the fluorescence intensity values of microarray spots corresponding to the biomarkers of interest, in addition to several experimental controls that verify correct functioning of the assay protocol. In this work, we report both dose-response characterization of the system using anti-E7 antibody introduced at known concentrations into serum and characterization of a set of clinical samples from which results were compared with a conventional enzyme-linked immunosorbent assay (ELISA) performed in microplate format. The demonstrated capability represents a simple, rapid, automated, and high-sensitivity method for multiplexed detection of protein biomarkers from a low-volume test sample.

  View details for DOI 10.1109/JSEN.2017.2777529

  View details for PubMedID 29881332

  View details for PubMedCentralID PMC5986186

• Guidance and Self-Sorting of Active Swimmers: 3D Periodic Arrays Increase Persistence Length of Human Sperm Selecting for the Fittest. Advanced science (Weinheim, Baden-Wurttemberg, Germany) Chinnasamy, T., Kingsley, J. L., Inci, F., Turek, P. J., Rosen, M. P., Behr, B., Tüzel, E., Demirci, U. 2018; 5 (2): 1700531

  Abstract

  Male infertility is a reproductive disease, and existing clinical solutions for this condition often involve long and cumbersome sperm sorting methods, including preprocessing and centrifugation-based steps. These methods also fall short when sorting for sperm free of reactive oxygen species, DNA damage, and epigenetic aberrations. Although several microfluidic platforms exist, they suffer from structural complexities, i.e., pumps or chemoattractants, setting insurmountable barriers to clinical adoption. Inspired by the natural filter-like capabilities of the female reproductive tract for sperm selection, a model-driven design, featuring pillar arrays that efficiently and noninvasively isolate highly motile and morphologically normal sperm, with lower epigenetic global methylation, from raw semen, is presented. The Simple Periodic ARray for Trapping And isolatioN (SPARTAN) created here modulates the directional persistence of sperm, increasing the spatial separation between progressive and nonprogressive motile sperm populations within an unprecedentedly short 10 min assay time. With over 99% motility of sorted sperm, a 5-fold improvement in morphology, 3-fold increase in nuclear maturity, and 2-4-fold enhancement in DNA integrity, SPARTAN offers to standardize sperm selection while eliminating operator-to-operator variations, centrifugation, and flow. SPARTAN can also be applied in other areas, including conservation ecology, breeding of farm animals, and design of flagellar microrobots for diagnostics.

  View details for DOI 10.1002/advs.201700531

  View details for PubMedID 29610725

  View details for PubMedCentralID PMC5827459

• An Automated Microfluidic Assay for Photonic Crystal Enhanced Detection and Analysis of an Antiviral Antibody Cancer Biomarker in Serum IEEE Sensors Journal Race, C. N. 2018; 18 (4): 1464 - 1473

  View details for DOI 10.1109/JSEN.2017.2777529

• Loss of Endothelial Derived WNT5A is Associated with Reduced Pericyte Recruitment and Small Vessel Loss in Pulmonary Arterial Hypertension. Circulation Yuan, K. n., Shamskhou, E. A., Orcholski, M. E., Nathan, A. n., Reddy, S. n., Honda, H. n., Mani, V. n., Zeng, Y. n., Ozen, M. O., Wang, L. n., Demirci, U. n., Tian, W. n., Nicolls, M. R., de Jesus Perez, V. A. 2018

  Abstract

  Pulmonary arterial hypertension (PAH) is a life-threatening disorder of the pulmonary circulation associated with loss and impaired regeneration of microvessels. Reduced pericyte coverage of pulmonary microvessels is a pathological feature of PAH and is partly due to the inability of pericytes to respond to signaling cues from neighboring pulmonary microvascular endothelial cells (PMVECs). We have shown that activation of the Wnt/PCP pathway is required for pericyte recruitment but whether production and release of specific Wnt ligands by PMVECs is responsible for Wnt/PCP activation in pericytes is unknown.Isolation of pericytes and PMVECs from healthy donor and PAH lungs was carried out using 3G5 or CD31 antibody conjugated magnetic beads. Wnt expression profile of PMVECs was documented via qPCR using a Wnt primer library. Exosome purification from PMVEC media was carried out using the ExoTIC device. Hemodynamic profile, right ventricular function and pulmonary vascular morphometry were obtained in a conditional endothelial specific Wnt5a knockout ( Wnt5aECKO) mouse model under normoxia, chronic hypoxia and hypoxia recovery.Quantification of Wnt ligand expression in healthy PMVECs co-cultured with pericytes demonstrated a 35-fold increase in Wnt5a, a known Wnt/PCP ligand. This Wnt5a spike was not seen in PAH PMVECs, which correlated with inability to recruit pericytes in matrigel co-culture assays. Exosomes purified from media demonstrated an increase in Wnt5a content when healthy PMVECs were co-cultured with pericytes, a finding that was not observed in exosomes of PAH PMVECs. Furthermore, the addition of either recombinant Wnt5a or purified healthy PMVEC exosomes increased pericyte recruitment to PAH PMVECs in co-culture studies. While no differences were noted in normoxia and chronic hypoxia, Wnt5aECKO mice demonstrated persistent pulmonary hypertension and right ventricular failure four weeks after recovery from chronic hypoxia, which correlated with significant reduction, muscularization and decreased pericyte coverage of microvessels.We identify Wnt5a as a key mediator for the establishment of pulmonary endothelial-pericyte interactions and its loss could contribute to PAH by reducing the viability of newly formed vessels. We speculate that therapies that mimic or restore Wnt5a production could help prevent loss of small vessels in PAH.

  View details for PubMedID 30586764

• Identification of hydrodynamic forces around 3D surrogates using particle image velocimetry in a microfluidic channel Afshar, S., Nath, S., Demirci, U., Hasan, T., Scarcelli, G., Rizvi, I., Franco, W., Gray, B. L., Becker, H. SPIE-INT SOC OPTICAL ENGINEERING. 2018

  View details for DOI 10.1117/12.2290372

  View details for Web of Science ID 000454729200002

• Monitoring Neutropenia for Cancer Patients at the Point of Care. Small methods Inan, H., Kingsley, J. L., Ozen, M. O., Tekin, H. C., Hoerner, C. R., Imae, Y., Metzner, T. J., Preiss, J. S., Durmus, N. G., Ozsoz, M., Wakelee, H., Fan, A. C., Tüzel, E., Demirci, U. 2017; 1 (9)

  Abstract

  Neutrophils have a critical role in regulating the immune system. The immune system is compromised during chemotherapy, increasing infection risks and imposing a need for regular monitoring of neutrophil counts. Although commercial hematology analyzers are currently used in clinical practice for neutrophil counts, they are only available in clinics and hospitals, use large blood volumes, and are not available at the point of care (POC). Additionally, phlebotomy and blood processing require trained personnel, where patients are often admitted to hospitals when the infections are at late stage due to lack of frequent monitoring. Here, a reliable method is presented that selectively captures and quantifies white blood cells (WBCs) and neutrophils from a finger prick volume of whole blood by integrating microfluidics with high-resolution imaging algorithms. The platform is compact, portable, and easy to use. It captures and quantifies WBCs and neutrophils with high efficiency (>95%) and specificity (>95%) with an overall 4.2% bias compared to standard testing. The results from a small cohort of patients (N = 11 healthy, N = 5 lung and kidney cancer) present a unique disposable cell counter, demonstrating the ability of this tool to monitor neutrophil and WBC counts within clinical or in resource-constrained environments.

  View details for DOI 10.1002/smtd.201700193

  View details for PubMedID 30740513

  View details for PubMedCentralID PMC6364993

• Bioacoustic-enabled patterning of human iPSC-derived cardiomyocytes into 3D cardiac tissue BIOMATERIALS Serpooshan, V., Chen, P., Wu, H., Lee, S., Sharma, A., Hu, D. A., Venkatraman, S., Ganesan, A. V., Usta, O. B., Yarmush, M., Yang, F., Wu, J. C., Demirci, U., Wu, S. M. 2017; 131: 47-57

  Abstract

  The creation of physiologically-relevant human cardiac tissue with defined cell structure and function is essential for a wide variety of therapeutic, diagnostic, and drug screening applications. Here we report a new scalable method using Faraday waves to enable rapid aggregation of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) into predefined 3D constructs. At packing densities that approximate native myocardium (10(8)-10(9) cells/ml), these hiPSC-CM-derived 3D tissues demonstrate significantly improved cell viability, metabolic activity, and intercellular connection when compared to constructs with random cell distribution. Moreover, the patterned hiPSC-CMs within the constructs exhibit significantly greater levels of contractile stress, beat frequency, and contraction-relaxation rates, suggesting their improved maturation. Our results demonstrate a novel application of Faraday waves to create stem cell-derived 3D cardiac tissue that resembles the cellular architecture of a native heart tissue for diverse basic research and clinical applications.

  View details for DOI 10.1016/j.biomaterials.2017.03.037

  View details for PubMedID 28376365

• Isolation, Detection, and Quantification of Cancer Biomarkers in HPV-Associated Malignancies. Scientific reports Inan, H., Wang, S., Inci, F., Baday, M., Zangar, R., Kesiraju, S., Anderson, K. S., Cunningham, B. T., Demirci, U. 2017; 7 (1): 3322

  Abstract

  Human Papillomavirus (HPV) infection has been recognized as the main etiologic factor in the development of various cancers including penile, vulva, oropharyngeal and cervical cancers. In the development of cancer, persistent HPV infections induce E6 and E7 oncoproteins, which promote cell proliferation and carcinogenesis resulting elevated levels of host antibodies (e.g., anti-HPV16 E7 antibody). Currently, these cancers are clinically diagnosed using invasive biopsy-based tests, which are performed only in centralized labs by experienced clinical staff using time-consuming and expensive tools and technologies. Therefore, these obstacles constrain their utilization at primary care clinics and in remote settings, where resources are limited. Here, we present a rapid, inexpensive, reliable, easy-to-use, customized immunoassay platform following a microfluidic filter device to detect and quantify anti-HPV16 E7 antibodies from whole blood as a non-invasive assisting technology for diagnosis of HPV-associated malignancies, especially, at primary healthcare and remote settings. The platform can detect and quantify anti-HPV16 E7 antibody down to 2.87 ng/mL. We further validated our immunoassay in clinical patient samples and it provided significantly high responses as compared to control samples. Thus, it can be potentially implemented as a pretesting tool to identify high-risk groups for broad monitoring of HPV-associated cancers in resource-constrained settings.

  View details for DOI 10.1038/s41598-017-02672-6

  View details for PubMedID 28607383

  View details for PubMedCentralID PMC5468352

• The promise of organ and tissue preservation to transform medicine. Nature biotechnology Giwa, S., Lewis, J. K., Alvarez, L., Langer, R., Roth, A. E., Church, G. M., Markmann, J. F., Sachs, D. H., Chandraker, A., Wertheim, J. A., Rothblatt, M., Boyden, E. S., Eidbo, E., Lee, W. P., Pomahac, B., Brandacher, G., Weinstock, D. M., Elliott, G., Nelson, D., Acker, J. P., Uygun, K., Schmalz, B., Weegman, B. P., Tocchio, A., Fahy, G. M., Storey, K. B., Rubinsky, B., Bischof, J., Elliott, J. A., Woodruff, T. K., Morris, G. J., Demirci, U., Brockbank, K. G., Woods, E. J., Ben, R. N., Baust, J. G., Gao, D., Fuller, B., Rabin, Y., Kravitz, D. C., Taylor, M. J., Toner, M. 2017; 35 (6): 530-542

  Abstract

  The ability to replace organs and tissues on demand could save or improve millions of lives each year globally and create public health benefits on par with curing cancer. Unmet needs for organ and tissue preservation place enormous logistical limitations on transplantation, regenerative medicine, drug discovery, and a variety of rapidly advancing areas spanning biomedicine. A growing coalition of researchers, clinicians, advocacy organizations, academic institutions, and other stakeholders has assembled to address the unmet need for preservation advances, outlining remaining challenges and identifying areas of underinvestment and untapped opportunities. Meanwhile, recent discoveries provide proofs of principle for breakthroughs in a family of research areas surrounding biopreservation. These developments indicate that a new paradigm, integrating multiple existing preservation approaches and new technologies that have flourished in the past 10 years, could transform preservation research. Capitalizing on these opportunities will require engagement across many research areas and stakeholder groups. A coordinated effort is needed to expedite preservation advances that can transform several areas of medicine and medical science.

  View details for DOI 10.1038/nbt.3889

  View details for PubMedID 28591112

• Bio-inspired Solute Enables Preservation of Human Oocytes using Minimum Volume Vitrification. Journal of tissue engineering and regenerative medicine Choi, J. K., El Assal, R., Ng, N., Ginsburg, E., Anchan, R. M., Demirci, U. 2017

  Abstract

  The ability to cryopreserve human oocytes has significant potential for fertility preservation. Current cryopreservation methods still suffer from the use of conventional cryoprotectants, such as dimethyl sulfoxide (DMSO), causing loss of viability and function. Such injuries result from the toxicity and high concentration of cryoprotectants as well as mechanical damage of cells due to ice crystal formation during the cooling and rewarming processes. Here, we report preservation of human oocytes following vitrification using an innovative bio-inspired cryoprotectant integrated with a minimum volume vitrification approach. The results demonstrate that the recovered human oocytes maintained viability following vitrification and rewarming. Moreover, when this approach was used to vitrify mouse oocytes, the recovered oocytes preserved their viability and function following vitrification and rewarming. This bio-inspired approach substitutes DMSO, a well-known toxic cryoprotectant, with ectoine, a non-toxic naturally occurring solute. The bio-inspired vitrification approach has potential to improve fertility preservation for women undergoing cancer treatment and endangered mammal species.

  View details for DOI 10.1002/term.2439

  View details for PubMedID 28481448

• High-throughput Characterization of HIV-1 Reservoir Reactivation Using a Single-Cell-in-Droplet PCR Assay. EBioMedicine Yucha, R. W., Hobbs, K. S., Hanhauser, E., Hogan, L. E., Nieves, W., Ozen, M. O., Inci, F., York, V., Gibson, E. A., Thanh, C., Shafiee, H., El Assal, R., Kiselinova, M., Robles, Y. P., Bae, H., Leadabrand, K. S., Wang, S., Deeks, S. G., Kuritzkes, D. R., Demirci, U., Henrich, T. J. 2017

  Abstract

  Reactivation of latent viral reservoirs is on the forefront of HIV-1 eradication research. However, it is unknown if latency reversing agents (LRAs) increase the level of viral transcription from cells producing HIV RNA or harboring transcriptionally-inactive (latent) infection. We therefore developed a microfluidic single-cell-in-droplet (scd)PCR assay to directly measure the number of CD4(+) T cells that produce unspliced (us)RNA and multiply spliced (ms)RNA following ex vivo latency reversal with either an histone deacetylase inhibitor (romidepsin) or T cell receptor (TCR) stimulation. Detection of HIV-1 transcriptional activity can also be performed on hundreds of thousands of CD4+ T-cells in a single experiment. The scdPCR method was then applied to CD4(+) T cells obtained from HIV-1-infected individuals on antiretroviral therapy. Overall, our results suggest that effects of LRAs on HIV-1 reactivation may be heterogeneous-increasing transcription from active cells in some cases and increasing the number of transcriptionally active cells in others. Genomic DNA and human mRNA isolated from HIV-1 reactivated cells could also be detected and quantified from individual cells. As a result, our assay has the potential to provide needed insight into various reservoir eradication strategies.

  View details for DOI 10.1016/j.ebiom.2017.05.006

  View details for PubMedID 28529033

• An integrated double-filtration microfluidic device for isolation, enrichment and quantification of urinary extracellular vesicles for detection of bladder cancer SCIENTIFIC REPORTS Liang, L., Kong, M., Zhou, S., Sheng, Y., Wang, P., Yu, T., Inci, F., Kuo, W. P., Li, L., Demirci, U., Wang, S. 2017; 7

  Abstract

  Extracellular vesicles (EVs), including exosomes and microvesicles, are present in a variety of bodily fluids, and the concentration of these sub-cellular vesicles and their associated biomarkers (proteins, nucleic acids, and lipids) can be used to aid clinical diagnosis. Although ultracentrifugation is commonly used for isolation of EVs, it is highly time-consuming, labor-intensive and instrument-dependent for both research laboratories and clinical settings. Here, we developed an integrated double-filtration microfluidic device that isolated and enriched EVs with a size range of 30-200 nm from urine, and subsequently quantified the EVs via a microchip ELISA. Our results showed that the concentration of urinary EVs was significantly elevated in bladder cancer patients (n = 16) compared to healthy controls (n = 8). Receiver operating characteristic (ROC) analysis demonstrated that this integrated EV double-filtration device had a sensitivity of 81.3% at a specificity of 90% (16 bladder cancer patients and 8 healthy controls). Thus, this integrated device has great potential to be used in conjunction with urine cytology and cystoscopy to improve clinical diagnosis of bladder cancer in clinics and at point-of-care (POC) settings.

  View details for DOI 10.1038/srep46224

  View details for Web of Science ID 000400049300001

  View details for PubMedID 28436447

• Photonic crystals: emerging biosensors and their promise for point-of-care applications. Chemical Society reviews Inan, H., Poyraz, M., Inci, F., Lifson, M. A., Baday, M., Cunningham, B. T., Demirci, U. 2017; 46 (2): 366-388

  Abstract

  Biosensors are extensively employed for diagnosing a broad array of diseases and disorders in clinical settings worldwide. The implementation of biosensors at the point-of-care (POC), such as at primary clinics or the bedside, faces impediments because they may require highly trained personnel, have long assay times, large sizes, and high instrumental cost. Thus, there exists a need to develop inexpensive, reliable, user-friendly, and compact biosensing systems at the POC. Biosensors incorporated with photonic crystal (PC) structures hold promise to address many of the aforementioned challenges facing the development of new POC diagnostics. Currently, PC-based biosensors have been employed for detecting a variety of biotargets, such as cells, pathogens, proteins, antibodies, and nucleic acids, with high efficiency and selectivity. In this review, we provide a broad overview of PCs by explaining their structures, fabrication techniques, and sensing principles. Furthermore, we discuss recent applications of PC-based biosensors incorporated with emerging technologies, including telemedicine, flexible and wearable sensing, smart materials and metamaterials. Finally, we discuss current challenges associated with existing biosensors, and provide an outlook for PC-based biosensors and their promise at the POC.

  View details for DOI 10.1039/c6cs00206d

  View details for PubMedID 27841420

• Paper-based analytical devices for clinical diagnosis: recent advances in the fabrication techniques and sensing mechanisms. Expert review of molecular diagnostics Sher, M., Zhuang, R., Demirci, U., Asghar, W. 2017

  Abstract

  There is a significant interest in developing inexpensive portable biosensing platforms for various applications including disease diagnostics, environmental monitoring, food safety, and water testing at the point-of-care (POC) settings. Current diagnostic assays available in the developed world require sophisticated laboratory infrastructure and expensive reagents. Hence, they are not suitable for resource-constrained settings with limited financial resources, basic health infrastructure, and few trained technicians. Cellulose and flexible transparency paper-based analytical devices have demonstrated enormous potential for developing robust, inexpensive and portable devices for disease diagnostics. These devices offer promising solutions to disease management in resource-constrained settings where the vast majority of the population cannot afford expensive and highly sophisticated treatment options. Areas covered: In this review, the authors describe currently developed cellulose and flexible transparency paper-based microfluidic devices, device fabrication techniques, and sensing technologies that are integrated with these devices. The authors also discuss the limitations and challenges associated with these devices and their potential in clinical settings. Expert commentary: In recent years, cellulose and flexible transparency paper-based microfluidic devices have demonstrated the potential to become future healthcare options despite a few limitations such as low sensitivity and reproducibility.

  View details for DOI 10.1080/14737159.2017.1285228

  View details for PubMedID 28103450

• Monitoring Neutropenia for Cancer Patients at the Point of Care Small Methods Inan, H., et al 2017

  View details for DOI 10.1002/smtd.201700193

• Tissue Engineering of 3D Organotypic Microtissues by Acoustic Assembly. Methods in molecular biology (Clifton, N.J.) Zhu, Y. n., Serpooshan, V. n., Wu, S. n., Demirci, U. n., Chen, P. n., Güven, S. n. 2017

  Abstract

  There is a rapidly growing interest in generation of 3D organotypic microtissues with human physiologically relevant structure, function, and cell population in a wide range of applications including drug screening, in vitro physiological/pathological models, and regenerative medicine. Here, we provide a detailed procedure to generate structurally defined 3D organotypic microtissues from cells or cell spheroids using acoustic waves as a biocompatible and scaffold-free tissue engineering tool.

  View details for PubMedID 28921421

• An Integrated Double-Filtration Microfluidic Device for Detection of Extracellular Vesicles from Urine for Bladder Cancer Diagnosis EXTRACELLULAR VESICLES: METHODS AND PROTOCOLS Liang, L., Sheng, Y., Zhou, S., Inci, F., Li, L., Demirci, U., Wang, S., Kuo, W. P., Jia, S. 2017; 1660: 355–64

  Abstract

  Extracellular vesicles (EVs) are present in a variety of bodily fluids and they play an important role in cellular communications and signal transduction mechanisms. Studies have shown that the number of EVs and EV-associated biomarkers (i.e., proteins, nucleic acids and lipids) can be used to aid clinical diagnosis. Although ultracentrifugation is commonly used for EV isolation, it is not practical for clinical settings. Here, we developed an integrated double-filtration device that isolated and enriched EVs from urine, and subsequently detected/quantified EVs from urine via microchip ELISA. Results showed that the concentration of EVs was significantly elevated compared to healthy controls. Receiver operating characteristic analysis demonstrated that this integrated EV quantification device had a sensitivity of 81.3% at a specificity of 90% (16 bladder cancer patients and eight healthy controls). Thus, this integrated device shows great potential to supplement urine cytology for diagnosis of bladder cancer in point-of-care (POC) settings.

  View details for PubMedID 28828671

• Guidance and Self-Sorting of Active Swimmers via 3-D Periodic Arrays Advanced Science Chinnasamy, T., Kingsley, J. L., Inci, F., Turek, P. J., Rosen, M. P., Behr, B., Tuzel, E., Demirci, U. 2017: 1700531

  Abstract

  Male infertility is a reproductive disease, and existing clinical solutions for this condition often involve long and cumbersome sperm sorting methods, including preprocessing and centrifugation-based steps. These methods also fall short when sorting for sperm free of reactive oxygen species, DNA damage, and epigenetic aberrations. Although several microfluidic platforms exist, they suffer from structural complexities, i.e., pumps or chemoattractants, setting insurmountable barriers to clinical adoption. Inspired by the natural filter-like capabilities of the female reproductive tract for sperm selection, a model-driven design, featuring pillar arrays that efficiently and noninvasively isolate highly motile and morphologically normal sperm, with lower epigenetic global methylation, from raw semen, is presented. The Simple Periodic ARray for Trapping And isolatioN (SPARTAN) created here modulates the directional persistence of sperm, increasing the spatial separation between progressive and nonprogressive motile sperm populations within an unprecedentedly short 10 min assay time. With over 99% motility of sorted sperm, a 5-fold improvement in morphology, 3-fold increase in nuclear maturity, and 2-4-fold enhancement in DNA integrity, SPARTAN offers to standardize sperm selection while eliminating operator-to-operator variations, centrifugation, and flow. SPARTAN can also be applied in other areas, including conservation ecology, breeding of farm animals, and design of flagellar microrobots for diagnostics.

  View details for DOI 10.1002/advs.201700531

  View details for PubMedCentralID PMC5827459

• Magnetically Guided Self-Assembly and Coding of 3D Living Architectures. Advanced materials (Deerfield Beach, Fla.) Tocchio, A. n., Durmus, N. G., Sridhar, K. n., Mani, V. n., Coskun, B. n., El Assal, R. n., Demirci, U. n. 2017

  Abstract

  In nature, cells self-assemble at the microscale into complex functional configurations. This mechanism is increasingly exploited to assemble biofidelic biological systems in vitro. However, precise coding of 3D multicellular living materials is challenging due to their architectural complexity and spatiotemporal heterogeneity. Therefore, there is an unmet need for an effective assembly method with deterministic control on the biomanufacturing of functional living systems, which can be used to model physiological and pathological behavior. Here, a universal system is presented for 3D assembly and coding of cells into complex living architectures. In this system, a gadolinium-based nonionic paramagnetic agent is used in conjunction with magnetic fields to levitate and assemble cells. Thus, living materials are fabricated with controlled geometry and organization and imaged in situ in real time, preserving viability and functional properties. The developed method provides an innovative direction to monitor and guide the reconfigurability of living materials temporally and spatially in 3D, which can enable the study of transient biological mechanisms. This platform offers broad applications in numerous fields, such as 3D bioprinting and bottom-up tissue engineering, as well as drug discovery, developmental biology, neuroscience, and cancer research.

  View details for PubMedID 29215164

• The Exosome Total Isolation Chip. ACS nano Liu, F. n., Vermesh, O. n., Mani, V. n., Ge, T. J., Madsen, S. J., Sabour, A. n., Hsu, E. C., Gowrishankar, G. n., Kanada, M. n., Jokerst, J. V., Sierra, R. G., Chang, E. n., Lau, K. n., Sridhar, K. n., Bermudez, A. n., Pitteri, S. J., Stoyanova, T. n., Sinclair, R. n., Nair, V. S., Gambhir, S. S., Demirci, U. n. 2017

  Abstract

  Circulating tumor-derived extracellular vesicles (EVs) have emerged as a promising source for identifying cancer biomarkers for early cancer detection. However, the clinical utility of EVs has thus far been limited by the fact that most EV isolation methods are tedious, nonstandardized, and require bulky instrumentation such as ultracentrifugation (UC). Here, we report a size-based EV isolation tool called ExoTIC (exosome total isolation chip), which is simple, easy-to-use, modular, and facilitates high-yield and high-purity EV isolation from biofluids. ExoTIC achieves an EV yield ∼4-1000-fold higher than that with UC, and EV-derived protein and microRNA levels are well-correlated between the two methods. Moreover, we demonstrate that ExoTIC is a modular platform that can sort a heterogeneous population of cancer cell line EVs based on size. Further, we utilize ExoTIC to isolate EVs from cancer patient clinical samples, including plasma, urine, and lavage, demonstrating the device's broad applicability to cancers and other diseases. Finally, the ability of ExoTIC to efficiently isolate EVs from small sample volumes opens up avenues for preclinical studies in small animal tumor models and for point-of-care EV-based clinical testing from fingerprick quantities (10-100 μL) of blood.

  View details for DOI 10.1021/acsnano.7b04878

  View details for PubMedID 29090896

• 3-D Microwell Array System for Culturing Virus Infected Tumor Cells SCIENTIFIC REPORTS El Assal, R., Gurkan, U. A., Chen, P., Juillard, F., Tocchio, A., Chinnasamy, T., Beauchemin, C., Unluisler, S., Canikyan, S., Holman, A., Srivatsa, S., Kaye, K. M., Demirci, U. 2016; 6

  Abstract

  Cancer cells have been increasingly grown in pharmaceutical research to understand tumorigenesis and develop new therapeutic drugs. Currently, cells are typically grown using two-dimensional (2-D) cell culture approaches, where the native tumor microenvironment is difficult to recapitulate. Thus, one of the main obstacles in oncology is the lack of proper infection models that recount main features present in tumors. In recent years, microtechnology-based platforms have been employed to generate three-dimensional (3-D) models that better mimic the native microenvironment in cell culture. Here, we present an innovative approach to culture Kaposi's sarcoma-associated herpesvirus (KSHV) infected human B cells in 3-D using a microwell array system. The results demonstrate that the KSHV-infected B cells can be grown up to 15 days in a 3-D culture. Compared with 2-D, cells grown in 3-D had increased numbers of KSHV latency-associated nuclear antigen (LANA) dots, as detected by immunofluorescence microscopy, indicating a higher viral genome copy number. Cells in 3-D also demonstrated a higher rate of lytic reactivation. The 3-D microwell array system has the potential to improve 3-D cell oncology models and allow for better-controlled studies for drug discovery.

  View details for DOI 10.1038/srep39144

  View details for Web of Science ID 000390304900001

  View details for PubMedID 28004818

  View details for PubMedCentralID PMC5177905

• Dynamic Microenvironment Induces Phenotypic Plasticity of Esophageal Cancer Cells Under Flow SCIENTIFIC REPORTS Kocal, G. C., Guven, S., Foygel, K., Goldman, A., Chen, P., Sengupta, S., Paulmurugan, R., Baskin, Y., Demirci, U. 2016; 6

  Abstract

  Cancer microenvironment is a remarkably heterogeneous composition of cellular and non-cellular components, regulated by both external and intrinsic physical and chemical stimuli. Physical alterations driven by increased proliferation of neoplastic cells and angiogenesis in the cancer microenvironment result in the exposure of the cancer cells to elevated levels of flow-based shear stress. We developed a dynamic microfluidic cell culture platform utilizing eshopagael cancer cells as model cells to investigate the phenotypic changes of cancer cells upon exposure to fluid shear stress. We report the epithelial to hybrid epithelial/mesenchymal transition as a result of decreasing E-Cadherin and increasing N-Cadherin and vimentin expressions, higher clonogenicity and ALDH positive expression of cancer cells cultured in a dynamic microfluidic chip under laminar flow compared to the static culture condition. We also sought regulation of chemotherapeutics in cancer microenvironment towards phenotypic control of cancer cells. Such in vitro microfluidic system could potentially be used to monitor how the interstitial fluid dynamics affect cancer microenvironment and plasticity on a simple, highly controllable and inexpensive bioengineered platform.

  View details for DOI 10.1038/srep38221

  View details for Web of Science ID 000389301200001

  View details for PubMedID 27910892

  View details for PubMedCentralID PMC5133540

• Quantification of Type, Timing, and Extent of Cell Body and Nucleus Deformations Caused by the Dimensions and Hydrophilicity of Square Prism Micropillars ADVANCED HEALTHCARE MATERIALS Hasturk, O., Sivas, A., Karasozen, B., Demirci, U., Hasirci, N., Hasirci, V. 2016; 5 (23): 2972-2982

  Abstract

  Novel digital analysis strategies are developed for the quantification of changes in the cytoskeletal and nuclear morphologies of mesenchymal stem cells cultured on micropillars. Severe deformations of nucleus and distinct conformational changes of cell body ranging from extensive elongation to branching are visualized and quantified. These deformations are caused mainly by the dimensions and hydrophilicity of the micropillars.

  View details for DOI 10.1002/adhm.201600857

  View details for Web of Science ID 000389920100002

  View details for PubMedID 27925459

• A high throughput approach for analysis of cell nuclear deformability at single cell level SCIENTIFIC REPORTS Ermis, M., Akkaynak, D., Chen, P., Demirci, U., Hasirci, V. 2016; 6

  Abstract

  Various physiological and pathological processes, such as cell differentiation, migration, attachment, and metastasis are highly dependent on nuclear elasticity. Nuclear morphology directly reflects the elasticity of the nucleus. We propose that quantification of changes in nuclear morphology on surfaces with defined topography will enable us to assess nuclear elasticity and deformability. Here, we used soft lithography techniques to produce 3 dimensional (3-D) cell culture substrates decorated with micron sized pillar structures of variable aspect ratios and dimensions to induce changes in cellular and nuclear morphology. We developed a high content image analysis algorithm to quantify changes in nuclear morphology at the single-cell level in response to physical cues from the 3-D culture substrate. We present that nuclear stiffness can be used as a physical parameter to evaluate cancer cells based on their lineage and in comparison to non-cancerous cells originating from the same tissue type. This methodology can be exploited for systematic study of mechanical characteristics of large cell populations complementing conventional tools such as atomic force microscopy and nanoindentation.

  View details for DOI 10.1038/srep36917

  View details for Web of Science ID 000387561900001

  View details for PubMedID 27841297

  View details for PubMedCentralID PMC5107983

• Flexible Substrate-Based Devices for Point-of-Care Diagnostics. Trends in biotechnology Wang, S., Chinnasamy, T., Lifson, M. A., Inci, F., Demirci, U. 2016; 34 (11): 909-921

  Abstract

  Point-of-care (POC) diagnostics play an important role in delivering healthcare, particularly for clinical management and disease surveillance in both developed and developing countries. Currently, the majority of POC diagnostics utilize paper substrates owing to affordability, disposability, and mass production capability. Recently, flexible polymer substrates have been investigated due to their enhanced physicochemical properties, potential to be integrated into wearable devices with wireless communications for personalized health monitoring, and ability to be customized for POC diagnostics. Here, we focus on the latest advances in developing flexible substrate-based diagnostic devices, including paper and polymers, and their clinical applications.

  View details for DOI 10.1016/j.tibtech.2016.05.009

  View details for PubMedID 27344425

• Microchip-based ultrafast serodiagnostic assay for tuberculosis SCIENTIFIC REPORTS Mani, V., Paleja, B., Larbi, K., Kumar, P., Tay, J. A., Siew, J. Y., Inci, F., Wang, S., Chee, C., Wang, Y. T., Demirci, U., De Libero, G., Singhal, A. 2016; 6

  Abstract

  Access to point-of-care (POC), rapid, inexpensive, sensitive, and instrument-free tests for the diagnosis of tuberculosis (TB) remains a major challenge. Here, we report a simple and low-cost microchip-based TB ELISA (MTBE) platform for the detection of anti-mycobacterial IgG in plasma samples in less than 15 minutes. The MTBE employs a flow-less, magnet-actuated, bead-based ELISA for simultaneous detection of IgG responses against multiple mycobacterial antigens. Anti-trehalose 6,6'-dimycolate (TDM) IgG responses were the strongest predictor for differentiating active tuberculosis (ATB) from healthy controls (HC) and latent tuberculosis infections (LTBI). The TDM-based MTBE demonstrated superior sensitivity compared to sputum microscopy (72% vs. 56%) with 80% and 63% positivity among smear-positive and smear-negative confirmed ATB samples, respectively. Receiver operating characteristic analysis indicated good accuracy for differentiating ATB from HC (AUC = 0.77). Thus, TDM-based MTBE can be potentially used as a screening device for rapid diagnosis of active TB at the POC.

  View details for DOI 10.1038/srep35845

  View details for Web of Science ID 000385927500001

  View details for PubMedID 27775039

  View details for PubMedCentralID PMC5075771

• Advances in biosensing strategies for HIV-1 detection, diagnosis, and therapeutic monitoring ADVANCED DRUG DELIVERY REVIEWS Lifson, M. A., Ozen, M. O., Inci, F., Wang, S., Inan, H., Baday, M., Henrich, T. J., Demirci, U. 2016; 103: 90-104

  Abstract

  HIV-1 is a major global epidemic that requires sophisticated clinical management. There have been remarkable efforts to develop new strategies for detecting and treating HIV-1, as it has been challenging to translate them into resource-limited settings. Significant research efforts have been recently devoted to developing point-of-care (POC) diagnostics that can monitor HIV-1 viral load with high sensitivity by leveraging micro- and nano-scale technologies. These POC devices can be applied to monitoring of antiretroviral therapy, during mother-to-child transmission, and identification of latent HIV-1 reservoirs. In this review, we discuss current challenges in HIV-1 diagnosis and therapy in resource-limited settings and present emerging technologies that aim to address these challenges using innovative solutions.

  View details for DOI 10.1016/j.addr.2016.05.018

  View details for Web of Science ID 000380083700007

  View details for PubMedID 27262924

  View details for PubMedCentralID PMC4943868

• Rapid Assembly of Heterogeneous 3D Cell Microenvironments in a Microgel Array ADVANCED MATERIALS Li, Y., Chen, P., Wang, Y., Yan, S., Feng, X., Du, W., Koehler, S. A., Demirci, U., Liu, B. 2016; 28 (18): 3543-?

  Abstract

  Heterogeneous 3D cell microenvironment arrays are rapidly assembled by combining surface-wettability-guided assembly and microdroplet-array-based operations. This approach enables precise control over individual shapes, sizes, chemical concentrations, cell density, and 3D spatial distribution of multiple components. This technique provides a cost-effective solution to meet the increasing demand of stem cell research, tissue engineering, and drug screening.

  View details for DOI 10.1002/adma.201600247

  View details for Web of Science ID 000376250600016

  View details for PubMedID 26991071

• Advances in addressing technical challenges of point-of-care diagnostics in resource-limited settings EXPERT REVIEW OF MOLECULAR DIAGNOSTICS Wang, S., Lifson, M. A., Inci, F., Liang, L., Sheng, Y., Demirci, U. 2016; 16 (4): 449-459

  View details for DOI 10.1586/14737159.2016.1142877

  View details for Web of Science ID 000372807200001

• Advances in addressing technical challenges of point-of-care diagnostics in resource-limited settings. Expert review of molecular diagnostics Wang, S., Lifson, M. A., Inci, F., Liang, L., Sheng, Y., Demirci, U. 2016; 16 (4): 449-459

  Abstract

  The striking prevalence of HIV, TB and malaria, as well as outbreaks of emerging infectious diseases, such as influenza A (H7N9), Ebola and MERS, poses great challenges for patient care in resource-limited settings (RLS). However, advanced diagnostic technologies cannot be implemented in RLS largely due to economic constraints. Simple and inexpensive point-of-care (POC) diagnostics, which rely less on environmental context and operator training, have thus been extensively studied to achieve early diagnosis and treatment monitoring in non-laboratory settings. Despite great input from material science, biomedical engineering and nanotechnology for developing POC diagnostics, significant technical challenges are yet to be overcome. Summarized here are the technical challenges associated with POC diagnostics from a RLS perspective and the latest advances in addressing these challenges are reviewed.

  View details for DOI 10.1586/14737159.2016.1142877

  View details for PubMedID 26777725

• Towards artificial tissue models: past, present, and future of 3D bioprinting BIOFABRICATION Arslan-Yildiz, A., El Assal, R., Chen, P., Guven, S., Inci, F., Demirci, U. 2016; 8 (1)

  View details for DOI 10.1088/1758-5090/8/1/014103

  View details for Web of Science ID 000373289000004

• Integrating Cell Phone Imaging with Magnetic Levitation (i-LEV) for Label-Free Blood Analysis at the Point-of-Living. Small Baday, M., Calamak, S., Durmus, N. G., Davis, R. W., Steinmetz, L. M., Demirci, U. 2016; 12 (9): 1222-1229

  Abstract

  There is an emerging need for portable, robust, inexpensive, and easy-to-use disease diagnosis and prognosis monitoring platforms to share health information at the point-of-living, including clinical and home settings. Recent advances in digital health technologies have improved early diagnosis, drug treatment, and personalized medicine. Smartphones with high-resolution cameras and high data processing power enable intriguing biomedical applications when integrated with diagnostic devices. Further, these devices have immense potential to contribute to public health in resource-limited settings where there is a particular need for portable, rapid, label-free, easy-to-use, and affordable biomedical devices to diagnose and continuously monitor patients for precision medicine, especially those suffering from rare diseases, such as sickle cell anemia, thalassemia, and chronic fatigue syndrome. Here, a magnetic levitation-based diagnosis system is presented in which different cell types (i.e., white and red blood cells) are levitated in a magnetic gradient and separated due to their unique densities. Moreover, an easy-to-use, smartphone incorporated levitation system for cell analysis is introduced. Using our portable imaging magnetic levitation (i-LEV) system, it is shown that white and red blood cells can be identified and cell numbers can be quantified without using any labels. In addition, cells levitated in i-LEV can be distinguished at single-cell resolution, potentially enabling diagnosis and monitoring, as well as clinical and research applications.

  View details for DOI 10.1002/smll.201501845

  View details for PubMedID 26523938

  View details for PubMedCentralID PMC4775401

• Engineering long shelf life multilayer biologically active surfaces on microfluidic devices for point of care applications SCIENTIFIC REPORTS Asghar, W., Yuksekkaya, M., Shafiee, H., Zhang, M., Ozen, M. O., Inci, F., Kocakulak, M., Demirci, U. 2016; 6

  Abstract

  Although materials and engineered surfaces are broadly utilized in creating assays and devices with wide applications in diagnostics, preservation of these immuno-functionalized surfaces on microfluidic devices remains a significant challenge to create reliable repeatable assays that would facilitate patient care in resource-constrained settings at the point-of-care (POC), where reliable electricity and refrigeration are lacking. To address this challenge, we present an innovative approach to stabilize surfaces on-chip with multiple layers of immunochemistry. The functionality of microfluidic devices using the presented method is evaluated at room temperature for up to 6-month shelf life. We integrated the preserved microfluidic devices with a lensless complementary metal oxide semiconductor (CMOS) imaging platform to count CD4(+) T cells from a drop of unprocessed whole blood targeting applications at the POC such as HIV management and monitoring. The developed immunochemistry stabilization method can potentially be applied broadly to other diagnostic immuno-assays such as viral load measurements, chemotherapy monitoring, and biomarker detection for cancer patients at the POC.

  View details for DOI 10.1038/srep21163

  View details for Web of Science ID 000370230000001

  View details for PubMedID 26883474

• Recapitulating cranial osteogenesis with neural crest cells in 3-D microenvironments. Acta biomaterialia Namkoong, B., Güven, S., Ramesan, S., Liaudanskaya, V., Abzhanov, A., Demirci, U. 2016; 31: 301-311

  Abstract

  The experimental systems that recapitulate the complexity of native tissues and enable precise control over the microenvironment are becoming essential for the pre-clinical tests of therapeutics and tissue engineering. Here, we described a strategy to develop an in vitro platform to study the developmental biology of craniofacial osteogenesis. In this study, we directly osteo-differentiated cranial neural crest cells (CNCCs) in a 3-D in vitro bioengineered microenvironment. Cells were encapsulated in the gelatin-based photo-crosslinkable hydrogel and cultured up to three weeks. We demonstrated that this platform allows efficient differentiation of p75 positive CNCCs to cells expressing osteogenic markers corresponding to the sequential developmental phases of intramembranous ossification. During the course of culture, we observed a decrease in the expression of early osteogenic marker Runx2, while the other mature osteoblast and osteocyte markers such as Osterix, Osteocalcin, Osteopontin and Bone sialoprotein increased. We analyzed the ossification of the secreted matrix with alkaline phosphatase and quantified the newly secreted hydroxyapatite. The Field Emission Scanning Electron Microscope (FESEM) images of the bioengineered hydrogel constructs revealed the native-like osteocytes, mature osteoblasts, and cranial bone tissue morphologies with canaliculus-like intercellular connections. This platform provides a broadly applicable model system to potentially study diseases involving primarily embryonic craniofacial bone disorders, where direct diagnosis and adequate animal disease models are limited.

  View details for DOI 10.1016/j.actbio.2015.12.004

  View details for PubMedID 26675129

• A Bio-Acoustic Levitational (BAL) Assembly Method for Engineering of Multilayered, 3D Brain-Like Constructs, Using Human Embryonic Stem Cell Derived Neuro-Progenitors ADVANCED MATERIALS Bouyer, C., Chen, P., Gueven, S., Demirtas, T. T., Nieland, T. J., Padilla, F., Demirci, U. 2016; 28 (1): 161-?

  View details for DOI 10.1002/adma.201503916

  View details for Web of Science ID 000367841100021

• A Bio-Acoustic Levitational (BAL) Assembly Method for Engineering of Multilayered, 3D Brain-Like Constructs, Using Human Embryonic Stem Cell Derived Neuro-Progenitors. Advanced materials (Deerfield Beach, Fla.) Bouyer, C., Chen, P., Güven, S., Demirtaş, T. T., Nieland, T. J., Padilla, F., Demirci, U. 2016; 28 (1): 161-7

  Abstract

  A bio-acoustic levitational assembly method for engineering of multilayered, 3D brainlike constructs is presented. Acoustic radiation forces are used to levitate neuroprogenitors derived from human embryonic stem cells in 3D multilayered fibrin tissue constructs. The neuro-progenitor cells are subsequently differentiated in neural cells, resulting in a 3D neuronal construct with inter and intralayer neurite elongations.

  View details for DOI 10.1002/adma.201503916

  View details for PubMedID 26554659

• Toxicology Study of Single-walled Carbon Nanotubes and Reduced Graphene Oxide in Human Sperm. Scientific reports Asghar, W., Shafiee, H., Velasco, V., Sah, V. R., Guo, S., El Assal, R., Inci, F., Rajagopalan, A., Jahangir, M., Anchan, R. M., Mutter, G. L., Ozkan, M., Ozkan, C. S., Demirci, U. 2016; 6: 30270-?

  Abstract

  Carbon-based nanomaterials such as single-walled carbon nanotubes and reduced graphene oxide are currently being evaluated for biomedical applications including in vivo drug delivery and tumor imaging. Several reports have studied the toxicity of carbon nanomaterials, but their effects on human male reproduction have not been fully examined. Additionally, it is not clear whether the nanomaterial exposure has any effect on sperm sorting procedures used in clinical settings. Here, we show that the presence of functionalized single walled carbon nanotubes (SWCNT-COOH) and reduced graphene oxide at concentrations of 1-25 μg/mL do not affect sperm viability. However, SWCNT-COOH generate significant reactive superoxide species at a higher concentration (25 μg/mL), while reduced graphene oxide does not initiate reactive species in human sperm. Further, we demonstrate that exposure to these nanomaterials does not hinder the sperm sorting process, and microfluidic sorting systems can select the sperm that show low oxidative stress post-exposure.

  View details for DOI 10.1038/srep30270

  View details for PubMedID 27538480

  View details for PubMedCentralID PMC4990966

• Engineering cancer microenvironments for in vitro 3-D tumor models MATERIALS TODAY Asghar, W., El Assal, R., Shafiee, H., Pitteri, S., Paulmurugan, R., Demirci, U. 2015; 18 (10): 539-553

  Abstract

  The natural microenvironment of tumors is composed of extracellular matrix (ECM), blood vasculature, and supporting stromal cells. The physical characteristics of ECM as well as the cellular components play a vital role in controlling cancer cell proliferation, apoptosis, metabolism, and differentiation. To mimic the tumor microenvironment outside the human body for drug testing, two-dimensional (2-D) and murine tumor models are routinely used. Although these conventional approaches are employed in preclinical studies, they still present challenges. For example, murine tumor models are expensive and difficult to adopt for routine drug screening. On the other hand, 2-D in vitro models are simple to perform, but they do not recapitulate natural tumor microenvironment, because they do not capture important three-dimensional (3-D) cell-cell, cell-matrix signaling pathways, and multi-cellular heterogeneous components of the tumor microenvironment such as stromal and immune cells. The three-dimensional (3-D) in vitro tumor models aim to closely mimic cancer microenvironments and have emerged as an alternative to routinely used methods for drug screening. Herein, we review recent advances in 3-D tumor model generation and highlight directions for future applications in drug testing.

  View details for DOI 10.1016/j.mattod.2015.05.002

  View details for Web of Science ID 000366576000019

  View details for PubMedCentralID PMC5407188

• 3-D tumor models. Materials today Asghar, W., El Assal, R., Shafiee, H., Pitteri, S., Paulmurugan, R., Demirci, U. 2015; 18 (10): 539-553

  Abstract

  The natural microenvironment of tumors is composed of extracellular matrix (ECM), blood vasculature, and supporting stromal cells. The physical characteristics of ECM as well as the cellular components play a vital role in controlling cancer cell proliferation, apoptosis, metabolism, and differentiation. To mimic the tumor microenvironment outside the human body for drug testing, two-dimensional (2-D) and murine tumor models are routinely used. Although these conventional approaches are employed in preclinical studies, they still present challenges. For example, murine tumor models are expensive and difficult to adopt for routine drug screening. On the other hand, 2-D in vitro models are simple to perform, but they do not recapitulate natural tumor microenvironment, because they do not capture important three-dimensional (3-D) cell-cell, cell-matrix signaling pathways, and multi-cellular heterogeneous components of the tumor microenvironment such as stromal and immune cells. The three-dimensional (3-D) in vitro tumor models aim to closely mimic cancer microenvironments and have emerged as an alternative to routinely used methods for drug screening. Herein, we review recent advances in 3-D tumor model generation and highlight directions for future applications in drug testing.

  View details for DOI 10.1016/j.mattod.2015.05.002

  View details for PubMedID 28458612

• Graphene-protein field effect biosensors: glucose sensing MATERIALS TODAY Viswanathan, S., Narayanan, T. N., Aran, K., Fink, K. D., Paredes, J., Ajayan, P. M., Filipek, S., Miszta, P., Tekin, H. C., Inci, F., Demirci, U., Li, P., Bolotin, K. I., Liepmann, D., Renugopalakrishanan, V. 2015; 18 (9): 513-522

  View details for DOI 10.1016/j.mattod.2015.04.003

  View details for Web of Science ID 000363533300018

• Portable lensless wide-field microscopy imaging platform based on digital inline holography and multi-frame pixel super-resolution. Light, science & applications Sobieranski, A. C., Inci, F., Tekin, H. C., Yuksekkaya, M., Comunello, E., Cobra, D., von Wangenheim, A., Demirci, U. 2015; 4

  Abstract

  In this paper, an irregular displacement-based lensless wide-field microscopy imaging platform is presented by combining digital in-line holography and computational pixel super-resolution using multi-frame processing. The samples are illuminated by a nearly coherent illumination system, where the hologram shadows are projected into a complementary metal-oxide semiconductor-based imaging sensor. To increase the resolution, a multi-frame pixel resolution approach is employed to produce a single holographic image from multiple frame observations of the scene, with small planar displacements. Displacements are resolved by a hybrid approach: (i) alignment of the LR images by a fast feature-based registration method, and (ii) fine adjustment of the sub-pixel information using a continuous optimization approach designed to find the global optimum solution. Numerical method for phase-retrieval is applied to decode the signal and reconstruct the morphological details of the analyzed sample. The presented approach was evaluated with various biological samples including sperm and platelets, whose dimensions are in the order of a few microns. The obtained results demonstrate a spatial resolution of 1.55 µm on a field-of-view of ≈30 mm2.

  View details for DOI 10.1038/lsa.2015.119

  View details for PubMedID 29657866

  View details for PubMedCentralID PMC5898403

• Deformation of a single mouse oocyte in a constricted microfluidic channel MICROFLUIDICS AND NANOFLUIDICS Luo, Z., Guven, S., Gozen, I., Chen, P., Tasoglu, S., Anchan, R. M., Bai, B., Demirci, U. 2015; 19 (4): 883-890

  View details for DOI 10.1007/s10404-015-1614-0

  View details for Web of Science ID 000361986400012

• Deformation of a single mouse oocyte in a constricted microfluidic channel. Microfluidics and nanofluidics Luo, Z., Guven, S., Gozen, I., Chen, P., Tasoglu, S., Anchan, R. M., Bai, B., Demirci, U. 2015; 19 (4): 883-890

  Abstract

  Single oocyte manipulation in microfluidic channels via precisely controlled flow is critical in microfluidic-based in vitro fertilization. Such systems can potentially minimize the number of transfer steps among containers for rinsing as often performed during conventional in vitro fertilization and can standardize protocols by minimizing manual handling steps. To study shape deformation of oocytes under shear flow and its subsequent impact on their spindle structure is essential for designing microfluidics for in vitro fertilization. Here, we developed a simple yet powerful approach to (i) trap a single oocyte and induce its deformation through a constricted microfluidic channel, (ii) quantify oocyte deformation in real-time using a conventional microscope, and (iii) retrieve the oocyte from the microfluidic device to evaluate changes in their spindle structures. We found that oocytes can be significantly deformed under high flow rates, e.g., 10 μl/min in a constricted channel with a width and height of 50 and 150 μm, respectively. Oocyte spindles can be severely damaged, as shown here by immunocytochemistry staining of the microtubules and chromosomes. The present approach can be useful to investigate underlying mechanisms of oocyte deformation exposed to well-controlled shear stresses in microfluidic channels, which enables a broad range of applications for reproductive medicine.

  View details for DOI 10.1007/s10404-015-1614-0

  View details for PubMedID 26696793

  View details for PubMedCentralID PMC4684828

• Portable lensless wide-field microscopy imaging platform based on digital inline holography and multi-frame pixel super-resolution LIGHT-SCIENCE & APPLICATIONS Sobieranski, A. C., Inci, F., Tekin, H. C., Yuksekkaya, M., Comunello, E., Cobra, D., von Wangenheim, A., Demirci, U. 2015; 4

  View details for DOI 10.1038/lsa.2015.119

  View details for Web of Science ID 000365013600006

• Biotunable Acoustic Node Assembly of Organoids ADVANCED HEALTHCARE MATERIALS Chen, P., Gueven, S., Usta, O. B., Yarmush, M. L., Demirci, U. 2015; 4 (13): 1937-1943

  View details for DOI 10.1002/adhm.201500279

  View details for Web of Science ID 000367752300003

• Biotunable Acoustic Node Assembly of Organoids. Advanced healthcare materials Chen, P., Güven, S., Usta, O. B., Yarmush, M. L., Demirci, U. 2015; 4 (13): 1937-1943

  Abstract

  Bioengineering of 3D microtissues from cell spheroids is demonstrated by employing the vibration of acoustic standing waves and its hydrodynamic effect at the bottom of a liquid-carrier chamber. A large number of cell spheroids (>10(4) ) are assembled in seconds into a closely packed structure in a scaffold-free fashion under nodal pattern of the standing waves in a fluidic environment.

  View details for DOI 10.1002/adhm.201500279

  View details for PubMedID 26149464

• Multitarget, quantitative nanoplasmonic electrical field-enhanced resonating device (NE2RD) for diagnostics. Proceedings of the National Academy of Sciences of the United States of America Inci, F., Filippini, C., Baday, M., Ozen, M. O., Calamak, S., Durmus, N. G., Wang, S., Hanhauser, E., Hobbs, K. S., Juillard, F., Kuang, P. P., Vetter, M. L., Carocci, M., Yamamoto, H. S., Takagi, Y., Yildiz, U. H., Akin, D., Wesemann, D. R., Singhal, A., Yang, P. L., Nibert, M. L., Fichorova, R. N., Lau, D. T., Henrich, T. J., Kaye, K. M., Schachter, S. C., Kuritzkes, D. R., Steinmetz, L. M., Gambhir, S. S., Davis, R. W., Demirci, U. 2015; 112 (32): E4354-63

  Abstract

  Recent advances in biosensing technologies present great potential for medical diagnostics, thus improving clinical decisions. However, creating a label-free general sensing platform capable of detecting multiple biotargets in various clinical specimens over a wide dynamic range, without lengthy sample-processing steps, remains a considerable challenge. In practice, these barriers prevent broad applications in clinics and at patients' homes. Here, we demonstrate the nanoplasmonic electrical field-enhanced resonating device (NE(2)RD), which addresses all these impediments on a single platform. The NE(2)RD employs an immunodetection assay to capture biotargets, and precisely measures spectral color changes by their wavelength and extinction intensity shifts in nanoparticles without prior sample labeling or preprocessing. We present through multiple examples, a label-free, quantitative, portable, multitarget platform by rapidly detecting various protein biomarkers, drugs, protein allergens, bacteria, eukaryotic cells, and distinct viruses. The linear dynamic range of NE(2)RD is five orders of magnitude broader than ELISA, with a sensitivity down to 400 fg/mL This range and sensitivity are achieved by self-assembling gold nanoparticles to generate hot spots on a 3D-oriented substrate for ultrasensitive measurements. We demonstrate that this precise platform handles multiple clinical samples such as whole blood, serum, and saliva without sample preprocessing under diverse conditions of temperature, pH, and ionic strength. The NE(2)RD's broad dynamic range, detection limit, and portability integrated with a disposable fluidic chip have broad applications, potentially enabling the transition toward precision medicine at the point-of-care or primary care settings and at patients' homes.

  View details for DOI 10.1073/pnas.1510824112

  View details for PubMedID 26195743

  View details for PubMedCentralID PMC4538635

• Multitarget, quantitative nanoplasmonic electrical field-enhanced resonating device ((NERD)-R-2) for diagnostics PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Inci, F., Filippini, C., Baday, M., Ozen, M. O., Calamak, S., Durmus, N. G., Wang, S., Hanhauser, E., Hobbs, K. S., Juillard, F., Kuang, P. P., Vetter, M. L., Carocci, M., Yamamoto, H. S., Takagi, Y., Yildiz, U. H., Akin, D., Wesemann, D. R., Singhal, A., Yang, P. L., Nibert, M. L., Fichorova, R. N., Lau, D. T., Henrich, T. J., Kaye, K. M., Schachter, S. C., Kuritzkes, D. R., Steinmetz, L. M., Gambhir, S. S., Davis, R. W., Demirci, U. 2015; 112 (32): E4354-E4363

  Abstract

  Recent advances in biosensing technologies present great potential for medical diagnostics, thus improving clinical decisions. However, creating a label-free general sensing platform capable of detecting multiple biotargets in various clinical specimens over a wide dynamic range, without lengthy sample-processing steps, remains a considerable challenge. In practice, these barriers prevent broad applications in clinics and at patients' homes. Here, we demonstrate the nanoplasmonic electrical field-enhanced resonating device (NE(2)RD), which addresses all these impediments on a single platform. The NE(2)RD employs an immunodetection assay to capture biotargets, and precisely measures spectral color changes by their wavelength and extinction intensity shifts in nanoparticles without prior sample labeling or preprocessing. We present through multiple examples, a label-free, quantitative, portable, multitarget platform by rapidly detecting various protein biomarkers, drugs, protein allergens, bacteria, eukaryotic cells, and distinct viruses. The linear dynamic range of NE(2)RD is five orders of magnitude broader than ELISA, with a sensitivity down to 400 fg/mL This range and sensitivity are achieved by self-assembling gold nanoparticles to generate hot spots on a 3D-oriented substrate for ultrasensitive measurements. We demonstrate that this precise platform handles multiple clinical samples such as whole blood, serum, and saliva without sample preprocessing under diverse conditions of temperature, pH, and ionic strength. The NE(2)RD's broad dynamic range, detection limit, and portability integrated with a disposable fluidic chip have broad applications, potentially enabling the transition toward precision medicine at the point-of-care or primary care settings and at patients' homes.

  View details for DOI 10.1073/pnas.1510824112

  View details for Web of Science ID 000359285100006

  View details for PubMedID 26195743

  View details for PubMedCentralID PMC4538635

• Hydrosoluble, UV-crosslinkable and injectable chitosan for patterned cell-laden microgel and rapid transdermal curing hydrogel in vivo ACTA BIOMATERIALIA Li, B., Wang, L., Xu, F., Gang, X., Demirci, U., Wei, D., Li, Y., Feng, Y., Jia, D., Zhou, Y. 2015; 22: 59-69

  Abstract

  Natural and biodegradable chitosan with unique amino groups has found widespread applications in tissue engineering and drug delivery. However, its applications have been limited by the poor solubility of native chitosan in neutral pH solution, which subsequently fails to achieve cell-laden hydrogel at physiological pH. To address this, we incorporated UV crosslinking ability in chitosan, allowing fabrication of patterned cell-laden and rapid transdermal curing hydrogel in vivo. The hydrosoluble, UV crosslinkable and injectable N-methacryloyl chitosan (N-MAC) was synthesized via single-step chemoselective N-acylation reaction, which simultaneously endowed chitosan with well solubility in neutral pH solution, UV crosslinkable ability and injectability. The solubility of N-MAC in neutral pH solution increased 2.21-fold with substitution degree increasing from 10.9% to 28.4%. The N-MAC allowed fabrication of cell-laden microgels with on-demand patterns via photolithography, and the cell viability in N-MAC hydrogel maintained 96.3 ± 1.3% N-MAC allowed rapid transdermal curing hydrogel in vivo within 60s through minimally invasive clinical surgery. Histological analysis revealed that low-dose UV irradiation hardly induced skin injury and acute inflammatory response disappeared after 7 days. N-MAC would allow rapid, robust and cost-effective fabrication of patterned cell-laden polysaccharide microgels with unique amino groups serving as building blocks for tissue engineering and rapid transdermal curing hydrogel in vivo for localized and sustained protein delivery.

  View details for DOI 10.1016/j.actbio.2015.04.026

  View details for Web of Science ID 000357241200007

  View details for PubMedID 25917845

• Magnetic Levitational Assembly for Living Material Fabrication ADVANCED HEALTHCARE MATERIALS Tasoglu, S., Yu, C. H., Liaudanskaya, V., Guven, S., Migliaresi, C., Demirci, U. 2015; 4 (10): 1469-1476

  Abstract

  Functional living materials with microscale compositional topographies are prevalent in nature. However, the creation of biomaterials composed of living micro building blocks, each programmed by composition, functionality, and shape, is still a challenge. A powerful yet simple approach to create living materials using a levitation-based magnetic method is presented.

  View details for DOI 10.1002/adhm.201500092

  View details for Web of Science ID 000358005500003

  View details for PubMedID 25872008

• Magnetic levitation of single cells. Proceedings of the National Academy of Sciences of the United States of America Durmus, N. G., Tekin, H. C., Guven, S., Sridhar, K., Arslan Yildiz, A., Calibasi, G., Ghiran, I., Davis, R. W., Steinmetz, L. M., Demirci, U. 2015; 112 (28): E3661-8

  Abstract

  Several cellular events cause permanent or transient changes in inherent magnetic and density properties of cells. Characterizing these changes in cell populations is crucial to understand cellular heterogeneity in cancer, immune response, infectious diseases, drug resistance, and evolution. Although magnetic levitation has previously been used for macroscale objects, its use in life sciences has been hindered by the inability to levitate microscale objects and by the toxicity of metal salts previously applied for levitation. Here, we use magnetic levitation principles for biological characterization and monitoring of cells and cellular events. We demonstrate that each cell type (i.e., cancer, blood, bacteria, and yeast) has a characteristic levitation profile, which we distinguish at an unprecedented resolution of 1 × 10(-4) g⋅mL(-1). We have identified unique differences in levitation and density blueprints between breast, esophageal, colorectal, and nonsmall cell lung cancer cell lines, as well as heterogeneity within these seemingly homogenous cell populations. Furthermore, we demonstrate that changes in cellular density and levitation profiles can be monitored in real time at single-cell resolution, allowing quantification of heterogeneous temporal responses of each cell to environmental stressors. These data establish density as a powerful biomarker for investigating living systems and their responses. Thereby, our method enables rapid, density-based imaging and profiling of single cells with intriguing applications, such as label-free identification and monitoring of heterogeneous biological changes under various physiological conditions, including antibiotic or cancer treatment in personalized medicine.

  View details for DOI 10.1073/pnas.1509250112

  View details for PubMedID 26124131

  View details for PubMedCentralID PMC4507238

• Magnetic levitation of single cells. Proceedings of the National Academy of Sciences of the United States of America Durmus, N. G., Tekin, H. C., Guven, S., Sridhar, K., Arslan Yildiz, A., Calibasi, G., Ghiran, I., Davis, R. W., Steinmetz, L. M., Demirci, U. 2015; 112 (28): E3661-8

  View details for DOI 10.1073/pnas.1509250112

  View details for PubMedID 26124131

• Levitational Image Cytometry with Temporal Resolution ADVANCED MATERIALS Tasoglu, S., Khoory, J. A., Tekin, H. C., Thomas, C., Karnoub, A. E., Ghiran, I. C., Demirci, U. 2015; 27 (26): 3901-?

  Abstract

  A simple, yet powerful magnetic-levitation-based device is reported for real-time, label-free separation, as well as high-resolution monitoring of cell populations based on their unique magnetic and density signatures. This method allows a wide variety of cellular processes to be studied, accompanied by transient or permanent changes in cells' fundamental characteristics as a biological material.

  View details for DOI 10.1002/adma.201405660

  View details for Web of Science ID 000357688900007

  View details for PubMedID 26058598

• Cytometry: Levitational Image Cytometry with Temporal Resolution (Adv. Mater. 26/2015). Advanced materials Tasoglu, S., Khoory, J. A., Tekin, H. C., Thomas, C., Karnoub, A. E., Ghiran, I. C., Demirci, U. 2015; 27 (26): 3900-?

  Abstract

  On page 3901, I. C. Ghiran, U. Demirci, and co-workers design a magnetic-levitation-based device that allows both label-free separation and high-resolution real-time monitoring of cell populations based on their unique magnetic and density signatures. This device can also be utilized to study transient or permanent changes in the fundamental characteristics of cells.

  View details for DOI 10.1002/adma.201570175

  View details for PubMedID 26149363

• Biomaterials: Magnetic Levitational Assembly for Living Material Fabrication (Adv. Healthcare Mater. 10/2015). Advanced healthcare materials Tasoglu, S., Yu, C. H., Liaudanskaya, V., Guven, S., Migliaresi, C., Demirci, U. 2015; 4 (10): 1420-?

  Abstract

  Functional living materials with microscale compositional topographies are prevalent in nature. However, the creation of biomaterials composed of living micro building blocks, each programmed by composition, functionality, and shape, is still a challenge. On page 1469, S. Tasoglu, U. Demirci, and co-workers present a powerful yet simple approach to create living materials using a levitation-based magnetic method.

  View details for DOI 10.1002/adhm.201570058

  View details for PubMedID 26173421

• Printed Flexible Plastic Microchip for Viral Load Measurement through Quantitative Detection of Viruses in Plasma and Saliva SCIENTIFIC REPORTS Shafiee, H., Kanakasabapathy, M. K., Juillard, F., Keser, M., Sadasivam, M., Yuksekkaya, M., Hanhauser, E., Henrich, T. J., Kuritzkes, D. R., Kaye, K. M., Demirci, U. 2015; 5

  Abstract

  We report a biosensing platform for viral load measurement through electrical sensing of viruses on a flexible plastic microchip with printed electrodes. Point-of-care (POC) viral load measurement is of paramount importance with significant impact on a broad range of applications, including infectious disease diagnostics and treatment monitoring specifically in resource-constrained settings. Here, we present a broadly applicable and inexpensive biosensing technology for accurate quantification of bioagents, including viruses in biological samples, such as plasma and artificial saliva, at clinically relevant concentrations. Our microchip fabrication is simple and mass-producible as we print microelectrodes on flexible plastic substrates using conductive inks. We evaluated the microchip technology by detecting and quantifying multiple Human Immunodeficiency Virus (HIV) subtypes (A, B, C, D, E, G, and panel), Epstein-Barr Virus (EBV), and Kaposi's Sarcoma-associated Herpes Virus (KSHV) in a fingerprick volume (50 µL) of PBS, plasma, and artificial saliva samples for a broad range of virus concentrations between 10(2) copies/mL and 10(7) copies/mL. We have also evaluated the microchip platform with discarded, de-identified HIV-infected patient samples by comparing our microchip viral load measurement results with reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) as the gold standard method using Bland-Altman Analysis.

  View details for DOI 10.1038/srep09919

  View details for Web of Science ID 000355859800001

  View details for PubMedID 26046668

  View details for PubMedCentralID PMC4456945

• Multiscale assembly for tissue engineering and regenerative medicine TRENDS IN BIOTECHNOLOGY Guven, S., Chen, P., Inci, F., Tasoglu, S., Erkmen, B., Demirci, U. 2015; 33 (5): 269-279

  Abstract

  Our understanding of cell biology and its integration with materials science has led to technological innovations in the bioengineering of tissue-mimicking grafts that can be utilized in clinical and pharmaceutical applications. Bioengineering of native-like multiscale building blocks provides refined control over the cellular microenvironment, thus enabling functional tissues. In this review, we focus on assembling building blocks from the biomolecular level to the millimeter scale. We also provide an overview of techniques for assembling molecules, cells, spheroids, and microgels and achieving bottom-up tissue engineering. Additionally, we discuss driving mechanisms for self- and guided assembly to create micro-to-macro scale tissue structures.

  View details for DOI 10.1016/j.tibtech.2015.02.003

  View details for Web of Science ID 000354157900005

  View details for PubMedID 25796488

• Portable Microfluidic Integrated Plasmonic Platform for Pathogen Detection SCIENTIFIC REPORTS Tokel, O., Yildiz, U. H., Inci, F., Durmus, N. G., Ekiz, O. O., Turker, B., Cetin, C., Rao, S., Sridhar, K., Natarajan, N., Shafiee, H., Dana, A., Demirci, U. 2015; 5

  Abstract

  Timely detection of infectious agents is critical in early diagnosis and treatment of infectious diseases. Conventional pathogen detection methods, such as enzyme linked immunosorbent assay (ELISA), culturing or polymerase chain reaction (PCR) require long assay times, and complex and expensive instruments, which are not adaptable to point-of-care (POC) needs at resource-constrained as well as primary care settings. Therefore, there is an unmet need to develop simple, rapid, and accurate methods for detection of pathogens at the POC. Here, we present a portable, multiplex, inexpensive microfluidic-integrated surface plasmon resonance (SPR) platform that detects and quantifies bacteria, i.e., Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) rapidly. The platform presented reliable capture and detection of E. coli at concentrations ranging from ~10(5) to 3.2 × 10(7) CFUs/mL in phosphate buffered saline (PBS) and peritoneal dialysis (PD) fluid. The multiplexing and specificity capability of the platform was also tested with S. aureus samples. The presented platform technology could potentially be applicable to capture and detect other pathogens at the POC and primary care settings.

  View details for DOI 10.1038/srep09152

  View details for Web of Science ID 000351699600001

  View details for PubMedID 25801042

  View details for PubMedCentralID PMC4371189

• Functional maintenance of differentiated embryoid bodies in microfluidic systems: a platform for personalized medicine. Stem cells translational medicine Guven, S., Lindsey, J. S., Poudel, I., Chinthala, S., Nickerson, M. D., Gerami-Naini, B., Gurkan, U. A., Anchan, R. M., Demirci, U. 2015; 4 (3): 261-268

  Abstract

  Hormone replacement therapies have become important for treating diseases such as premature ovarian failure or menopausal complications. The clinical use of bioidentical hormones might significantly reduce some of the potential risks reportedly associated with the use of synthetic hormones. In the present study, we demonstrate the utility and advantage of a microfluidic chip culture system to enhance the development of personalized, on-demand, treatment modules using embryoid bodies (EBs). Functional EBs cultured on microfluidic chips represent a platform for personalized, patient-specific treatment cassettes that can be cryopreserved until required for treatment. We assessed the viability, differentiation, and functionality of EBs cultured and cryopreserved in this system. During extended microfluidic culture, estradiol, progesterone, testosterone, and anti-müllerian hormone levels were measured, and the expression of differentiated steroidogenic cells was confirmed by immunocytochemistry assay for the ovarian tissue markers anti-müllerian hormone receptor type II, follicle-stimulating hormone receptor, and inhibin β-A and the estrogen biosynthesis enzyme aromatase. Our studies showed that under microfluidic conditions, differentiated steroidogenic EBs continued to secrete estradiol and progesterone at physiologically relevant concentrations (30-120 pg/ml and 150-450 pg/ml, respectively) for up to 21 days. Collectively, we have demonstrated for the first time the feasibility of using a microfluidic chip system with continuous flow for the differentiation and extended culture of functional steroidogenic stem cell-derived EBs, the differentiation of EBs into cells expressing ovarian antigens in a microfluidic system, and the ability to cryopreserve this system with restoration of growth and functionality on thawing. These results present a platform for the development of a new therapeutic system for personalized medicine.

  View details for DOI 10.5966/sctm.2014-0119

  View details for PubMedID 25666845

  View details for PubMedCentralID PMC4339847

• Highlights from the latest articles in advanced biomanufacturing at micro- and nano-scale. Nanomedicine Assal, R. E., Chen, P., Demirci, U. 2015; 10 (3): 347-350

  View details for DOI 10.2217/nnm.14.210

  View details for PubMedID 25707972

• Advances in Nanotechnology and Microfluidics for Human Papillomavirus Diagnostics PROCEEDINGS OF THE IEEE Tasoglu, S., Tekin, H. C., Inci, F., Knowlton, S., Wang, S., Wang-Johanning, F., Johanning, G., Colevas, D., Demirci, U. 2015; 103 (2): 161-178

  View details for DOI 10.1109/JPROC.2014.2384836

  View details for Web of Science ID 000352156900004

• Emerging Technologies for Point-of-Care Management of HIV Infection ANNUAL REVIEW OF MEDICINE, VOL 66 Shafiee, H., Wang, S., Inci, F., Toy, M., Henrich, T. J., Kuritzkes, D. R., Demirci, U. 2015; 66: 387-405

  Abstract

  The global HIV/AIDS pandemic has resulted in 39 million deaths to date, and there are currently more than 35 million people living with HIV worldwide. Prevention, screening, and treatment strategies have led to major progress in addressing this disease globally. Diagnostics is critical for HIV prevention, screening and disease staging, and monitoring antiretroviral therapy (ART). Currently available diagnostic assays, which include polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and western blot (WB), are complex, expensive, and time consuming. These diagnostic technologies are ill suited for use in low- and middle-income countries, where the challenge of the HIV/AIDS pandemic is most severe. Therefore, innovative, inexpensive, disposable, and rapid diagnostic platform technologies are urgently needed. In this review, we discuss challenges associated with HIV management in resource-constrained settings and review the state-of-the-art HIV diagnostic technologies for CD4(+) T lymphocyte count, viral load measurement, and drug resistance testing.

  View details for DOI 10.1146/annurev-med-092112-143017

  View details for Web of Science ID 000348560300026

  View details for PubMedID 25423597

• Flexible Microwave Antenna Applicator for Chemo-Thermotherapy of the Breast IEEE ANTENNAS AND WIRELESS PROPAGATION LETTERS Asili, M., Chen, P., Hood, A. Z., Purser, A., Hulsey, R., Johnson, L., Ganesan, A. V., Demirci, U., Topsakal, E. 2015; 14: 1778-1781

  View details for DOI 10.1109/LAWP.2015.2423655

  View details for Web of Science ID 000362007200004

• Paper and flexible substrates as materials for biosensing platforms to detect multiple biotargets. Scientific reports Shafiee, H., Asghar, W., Inci, F., Yuksekkaya, M., Jahangir, M., Zhang, M. H., Durmus, N. G., Gurkan, U. A., Kuritzkes, D. R., Demirci, U. 2015; 5: 8719-?

  Abstract

  The need for sensitive, robust, portable, and inexpensive biosensing platforms is of significant interest in clinical applications for disease diagnosis and treatment monitoring at the point-of-care (POC) settings. Rapid, accurate POC diagnostic assays play a crucial role in developing countries, where there are limited laboratory infrastructure, trained personnel, and financial support. However, current diagnostic assays commonly require long assay time, sophisticated infrastructure and expensive reagents that are not compatible with resource-constrained settings. Although paper and flexible material-based platform technologies provide alternative approaches to develop POC diagnostic assays for broad applications in medicine, they have technical challenges integrating to different detection modalities. Here, we address the limited capability of current paper and flexible material-based platforms by integrating cellulose paper and flexible polyester films as diagnostic biosensing materials with various detection modalities through the development and validation of new widely applicable electrical and optical sensing mechanisms using antibodies and peptides. By incorporating these different detection modalities, we present selective and accurate capture and detection of multiple biotargets including viruses (Human Immunodeficieny Virus-1), bacteria (Escherichia coli and Staphylococcus aureus), and cells (CD4(+) T lymphocytes) from fingerprick volume equivalent of multiple biological specimens such as whole blood, plasma, and peritoneal dialysis effluent with clinically relevant detection and sensitivity.

  View details for DOI 10.1038/srep08719

  View details for PubMedID 25743880

  View details for PubMedCentralID PMC4351531

• Recent advances in micro/nanotechnologies for global control of hepatitis B infection BIOTECHNOLOGY ADVANCES Yildiz, U. H., Inci, F., Wang, S., Toy, M., Tekin, H. C., Javaid, A., Lau, D. T., Demirci, U. 2015; 33 (1): 178-190

  Abstract

  The control of hepatitis B virus (HBV) infection is a challenging task, specifically in developing countries there is limited access to diagnostics and antiviral treatment mainly due to high costs and insufficient healthcare infrastructure. Although the current diagnostic technologies can reliably detect HBV, they are relatively laborious, impractical and require expensive resources that are not suitable for resource-limited settings. Advances in micro/nanotechnology are pioneering the development of new generation methodologies in diagnosis and screening of HBV. Owing to combination of nanomaterials (metal/inorganic nanoparticles, carbon nanotubes, etc.) with microfabrication technologies, utilization of miniaturized sensors detecting HBV and other viruses from ultra-low volume of blood, serum and plasma is realized. The state-of-the-art microfluidic devices with integrated nanotechnologies potentially allow for inexpensive HBV screening at low cost. This review aims to highlight recent advances in nanotechnology and microfabrication processes that are employed for developing point-of-care (POC) HBV assays.

  View details for DOI 10.1016/j.biotechadv.2014.11.003

  View details for Web of Science ID 000351321400013

  View details for PubMedID 25450190

• Microchip ELISA Coupled with Cell Phone to Detect Ovarian Cancer HE4 Biomarker in Urine. Methods in molecular biology (Clifton, N.J.) Wang, S., Akbas, R., Demirci, U. 2015; 1256: 111-121

  Abstract

  Ovarian cancer is a leading cause of death from gynecologic cancers in the USA, and early diagnosis can potentially increase 5-year survival rate. Detection of biomarkers derived from hyperplasia of epithelial tissue by enzyme-linked immunosorbent assay (ELISA) proves to be a practical way of early diagnosis of ovarian cancer. However, ELISA is commonly performed in a laboratory setting, and it cannot be used in a clinical setting for on-site consultation. We have shown a microchip ELISA that detects HE4, an ovarian cancer biomarker, from urine using a cell phone integrated with a mobile application for imaging and data analysis. In microchip ELISA, HE4 from urine was first absorbed on the surface; the primary and secondary antibodies were subsequently anchored on the surface via immuno-reaction; and addition of substrate led to color development because of enzymatic labeling. The microchip after color development was imaged using a cell phone, and the color intensity was analyzed by an integrated mobile application. By comparing with an ELISA standard curve, the concentration of HE4 was reported on the cell phone screen. The presented microchip ELISA coupled with a cell phone is portable as opposed to traditional ELISA, and this method can facilitate the detection of ovarian cancer at the point-of-care (POC).

  View details for DOI 10.1007/978-1-4939-2172-0_8

  View details for PubMedID 25626535

• Emerging technologies for monitoring drug-resistant tuberculosis at the point-of-care ADVANCED DRUG DELIVERY REVIEWS Mani, V., Wang, S., Inci, F., De Libero, G., Singhal, A., Demirci, U. 2014; 78: 105-117

  Abstract

  Infectious diseases are the leading cause of death worldwide. Among them, tuberculosis (TB) remains a major threat to public health, exacerbated by the emergence of multiple drug-resistant (MDR) and extensively drug-resistant (XDR) Mycobacterium tuberculosis (Mtb). MDR-Mtb strains are resistant to first-line anti-TB drugs such as isoniazid and rifampicin; whereas XDR-Mtb strains are resistant to additional drugs including at least to any fluoroquinolone and one of the second-line anti-TB injectable drugs such as kanamycin, capreomycin, or amikacin. Clinically, these strains have significantly impacted the management of TB in high-incidence developing countries, where systemic surveillance of TB drug resistance is lacking. For effective management of TB on-site, early detection of drug resistance is critical to initiate treatment, to reduce mortality, and to thwart drug-resistant TB transmission. In this review, we discuss the diagnostic challenges to detect drug-resistant TB at the point-of-care (POC). Moreover, we present the latest advances in nano/microscale technologies that can potentially detect TB drug resistance to improve on-site patient care.

  View details for DOI 10.1016/j.addr.2014.05.015

  View details for Web of Science ID 000358460400009

  View details for PubMedCentralID PMC4254374

• Emerging technologies for monitoring drug-resistant tuberculosis at the point-of-care. Advanced drug delivery reviews Mani, V., Wang, S., Inci, F., De Libero, G., Singhal, A., Demirci, U. 2014; 78: 105-17

  Abstract

  Infectious diseases are the leading cause of death worldwide. Among them, tuberculosis (TB) remains a major threat to public health, exacerbated by the emergence of multiple drug-resistant (MDR) and extensively drug-resistant (XDR) Mycobacterium tuberculosis (Mtb). MDR-Mtb strains are resistant to first-line anti-TB drugs such as isoniazid and rifampicin; whereas XDR-Mtb strains are resistant to additional drugs including at least to any fluoroquinolone and one of the second-line anti-TB injectable drugs such as kanamycin, capreomycin, or amikacin. Clinically, these strains have significantly impacted the management of TB in high-incidence developing countries, where systemic surveillance of TB drug resistance is lacking. For effective management of TB on-site, early detection of drug resistance is critical to initiate treatment, to reduce mortality, and to thwart drug-resistant TB transmission. In this review, we discuss the diagnostic challenges to detect drug-resistant TB at the point-of-care (POC). Moreover, we present the latest advances in nano/microscale technologies that can potentially detect TB drug resistance to improve on-site patient care.

  View details for DOI 10.1016/j.addr.2014.05.015

  View details for PubMedID 24882226

  View details for PubMedCentralID PMC4254374

• Two-dimensional numerical study of flow dynamics of a nucleated cell tethered under shear flow CHEMICAL ENGINEERING SCIENCE Luo, Z. Y., He, L., Wang, S. Q., Tasoglu, S., Xu, F., Demirci, U., Bai, B. F. 2014; 119: 236-244

  View details for DOI 10.1016/j.ces.2014.07.048

  View details for Web of Science ID 000342680700024

• Selection of Functional Human Sperm with Higher DNA Integrity and Fewer Reactive Oxygen Species ADVANCED HEALTHCARE MATERIALS Asghar, W., Velasco, V., Kingsley, J. L., Shoukat, M. S., Shafiee, H., Anchan, R. M., Mutter, G. L., Tuezel, E., Demirci, U. 2014; 3 (10): 1671-1679

  View details for DOI 10.1002/adhm.201400058

  View details for Web of Science ID 000343798800016

• Nanomechanical motion of Escherichia coli adhered to a surface APPLIED PHYSICS LETTERS Lissandrello, C., Inci, F., Francom, M., Paul, M. R., Demirci, U., Ekinci, K. L. 2014; 105 (11)

  View details for DOI 10.1063/1.4895132

  View details for Web of Science ID 000342995800101

• Microscale Assembly Directed by Liquid-Based Template ADVANCED MATERIALS Chen, P., Luo, Z., Gueven, S., Tasoglu, S., Ganesan, A. V., Weng, A., Demirci, U. 2014; 26 (34): 5936-?

  View details for DOI 10.1002/adma.201402079

  View details for Web of Science ID 000342147400004

• Microscale assembly directed by liquid-based template. Advanced materials (Deerfield Beach, Fla.) Chen, P., Luo, Z., Güven, S., Tasoglu, S., Ganesan, A. V., Weng, A., Demirci, U. 2014; 26 (34): 5936-41

  Abstract

  A liquid surface established by standing waves is used as a dynamically reconfigurable template to assemble microscale materials into ordered, symmetric structures in a scalable and parallel manner. The broad applicability of this technology is illustrated by assembling diverse materials from soft matter, rigid bodies, individual cells, cell spheroids and cell-seeded microcarrier beads.

  View details for DOI 10.1002/adma.201402079

  View details for PubMedID 24956442

  View details for PubMedCentralID PMC4159433

• Bio-inspired cryo-ink preserves red blood cell phenotype and function during nanoliter vitrification. Advanced materials El Assal, R., Guven, S., Gurkan, U. A., Gozen, I., Shafiee, H., Dalbeyler, S., Abdalla, N., Thomas, G., Fuld, W., Illigens, B. M., Estanislau, J., Khoory, J., Kaufman, R., Zylberberg, C., Lindeman, N., Wen, Q., Ghiran, I., Demirci, U. 2014; 26 (33): 5815-5822

  Abstract

  Current red-blood-cell cryopreservation methods utilize bulk volumes, causing cryo-injury of cells, which results in irreversible disruption of cell morphology, mechanics, and function. An innovative approach to preserve human red-blood-cell morphology, mechanics, and function following vitrification in nanoliter volumes is developed using a novel cryo-ink integrated with a bioprinting approach.

  View details for DOI 10.1002/adma.201400941

  View details for PubMedID 25047246

• Preserving human cells for regenerative, reproductive, and transfusion medicine. Biotechnology journal Asghar, W., El Assal, R., Shafiee, H., Anchan, R. M., Demirci, U. 2014; 9 (7): 895-903

  Abstract

  Cell cryopreservation maintains cellular life at sub-zero temperatures by slowing down biochemical processes. Various cell types are routinely cryopreserved in modern reproductive, regenerative, and transfusion medicine. Current cell cryopreservation methods involve freezing (slow/rapid) or vitrifying cells in the presence of a cryoprotective agent (CPA). Although these methods are clinically utilized, cryo-injury due to ice crystals, osmotic shock, and CPA toxicity cause loss of cell viability and function. Recent approaches using minimum volume vitrification provide alternatives to the conventional cryopreservation methods. Minimum volume vitrification provides ultra-high cooling and rewarming rates that enable preserving cells without ice crystal formation. Herein, we review recent advances in cell cryopreservation technology and provide examples of techniques that are utilized in oocyte, stem cell, and red blood cell cryopreservation.

  View details for DOI 10.1002/biot.201300074

  View details for PubMedID 24995723

• Engineering Anisotropic Biomimetic Fibrocartilage Microenvironment by Bioprinting Mesenchymal Stem Cells in Nanoliter Gel Droplets MOLECULAR PHARMACEUTICS Gurkan, U. A., El Assal, R., Yildiz, S. E., Sung, Y., Trachtenberg, A. J., Kuo, W. P., Demirci, U. 2014; 11 (7): 2151-2159

  Abstract

  Over the past decade, bioprinting has emerged as a promising patterning strategy to organize cells and extracellular components both in two and three dimensions (2D and 3D) to engineer functional tissue mimicking constructs. So far, tissue printing has neither been used for 3D patterning of mesenchymal stem cells (MSCs) in multiphase growth factor embedded 3D hydrogels nor been investigated phenotypically in terms of simultaneous differentiation into different cell types within the same micropatterned 3D tissue constructs. Accordingly, we demonstrated a biochemical gradient by bioprinting nanoliter droplets encapsulating human MSCs, bone morphogenetic protein 2 (BMP-2), and transforming growth factor β1 (TGF- β1), engineering an anisotropic biomimetic fibrocartilage microenvironment. Assessment of the model tissue construct displayed multiphasic anisotropy of the incorporated biochemical factors after patterning. Quantitative real time polymerase chain reaction (qRT-PCR) results suggested genomic expression patterns leading to simultaneous differentiation of MSC populations into osteogenic and chondrogenic phenotype within the multiphasic construct, evidenced by upregulation of osteogenesis and condrogenesis related genes during in vitro culture. Comprehensive phenotypic network and pathway analysis results, which were based on genomic expression data, indicated activation of differentiation related mechanisms, via signaling pathways, including TGF, BMP, and vascular endothelial growth factor.

  View details for DOI 10.1021/mp400573g

  View details for Web of Science ID 000338748200024

  View details for PubMedID 24495169

• Evaluation of Epithelial Chimerism After Bone Marrow Mesenchymal Stromal Cell Infusion in Intestinal Transplant Patients 13th International Small Bowel Transplant Symposium Kilinc, S., Gurkan, U. A., Guven, S., Koyuncu, G., Tan, S., Karaca, C., Ozdogan, O., Dogan, M., Tugmen, C., Pala, E. E., Bayol, U., Baran, M., Kurtulmus, Y., Pirim, I., Kebapci, E., Demirci, U. ELSEVIER SCIENCE INC. 2014: 2125–32

  Abstract

  Intestinal transplantation is the most effective treatment for patients with short bowel syndrome and small bowel insufficiencies. We evaluated epithelial chimerism after infusion of autologous bone marrow mesenchymal stromal cells (BMSCs) in patients undergoing cadaveric donor isolated intestinal transplantation (I-ITx). BMSCs were isolated from patients' bone marrow via iliac puncture and expanded in vitro prior to infusion. Two out of the 3 patients were infused with autologous BMSCs, and small intestine tissue biopsies collected post-operatively were analyzed for epithelial chimerism using XY fluorescent in situ hybridization and short tandem repeat polymerase chain reaction. We observed epithelial chimeric effect in conditions both with and without BMSC infusion. Although our results suggest a higher epithelial chimerism effect with autologous BMSC infusion in I-ITx, the measurements in multiple biopsies at different time points that demonstrate the reproducibility of this finding and its stability or changes in the level over time would be beneficial. These approaches may have potential implications for improved graft survival, lower immunosuppressant doses, superior engraftment of the transplanted tissue, and higher success rates in I-ITx.

  View details for DOI 10.1016/j.transproceed.2014.06.039

  View details for Web of Science ID 000341076800117

  View details for PubMedID 25131122

• Advances in Plasmonic Technologies for Point of Care Applications CHEMICAL REVIEWS Tokel, O., Inci, F., Demirci, U. 2014; 114 (11): 5728-5752

  View details for DOI 10.1021/cr4000623

  View details for Web of Science ID 000337336500004

• Nanostructured Optical Photonic Crystal Biosensor for HIV Viral Load Measurement SCIENTIFIC REPORTS Shafiee, H., Lidstone, E. A., Jahangir, M., Inci, F., Hanhauser, E., Henrich, T. J., Kuritzkes, D. R., Cunningham, B. T., Demirci, U. 2014; 4

  Abstract

  Detecting and quantifying biomarkers and viruses in biological samples have broad applications in early disease diagnosis and treatment monitoring. We have demonstrated a label-free optical sensing mechanism using nanostructured photonic crystals (PC) to capture and quantify intact viruses (HIV-1) from biologically relevant samples. The nanostructured surface of the PC biosensor resonantly reflects a narrow wavelength band during illumination with a broadband light source. Surface-adsorbed biotarget induces a shift in the resonant Peak Wavelength Value (PWV) that is detectable with <10 pm wavelength resolution, enabling detection of both biomolecular layers and small number of viruses that sparsely populate the transducer surface. We have successfully captured and detected HIV-1 in serum and phosphate buffered saline (PBS) samples with viral loads ranging from 10(4) to 10(8) copies/mL. The surface density of immobilized biomolecular layers used in the sensor functionalization process, including 3-mercaptopropyltrimethoxysilane (3-MPS), N-gamma-Maleimidobutyryl-oxysuccinimide ester (GMBS), NeutrAvidin, anti-gp120, and bovine serum albumin (BSA) were also quantified by the PC biosensor.

  View details for DOI 10.1038/srep04116

  View details for Web of Science ID 000332014100001

  View details for PubMedID 24576941

• Use of commercial off-the-shelf digital cameras for scientific data acquisition and scene-specific color calibration JOURNAL OF THE OPTICAL SOCIETY OF AMERICA A-OPTICS IMAGE SCIENCE AND VISION Akkaynak, D., Treibitz, T., Xiao, B., Guerkan, U. A., Allen, J. J., Demirci, U., Hanlon, R. T. 2014; 31 (2): 312-321

  View details for DOI 10.1364/JOSAA.31.000312

  View details for Web of Science ID 000331070600013

• Micro-a-fluidics ELISA for Rapid CD4 Cell Count at the Point-of-Care SCIENTIFIC REPORTS Wang, S., Tasoglu, S., Chen, P. Z., Chen, M., Akbas, R., Wach, S., Ozdemir, C. I., Gurkan, U. A., Giguel, F. F., Kuritzkes, D. R., Demirci, U. 2014; 4

  Abstract

  HIV has become one of the most devastating pathogens in human history. Despite fast progress in HIV-related basic research, antiretroviral therapy (ART) remains the most effective method to save AIDS patients' lives. Unfortunately, ART cannot be universally accessed, especially in developing countries, due to the lack of effective treatment monitoring diagnostics. Here, we present an inexpensive, rapid and portable micro-a-fluidic platform, which can streamline the process of an enzyme-linked immunosorbent assay (ELISA) in a fully automated manner for CD4 cell count. The micro-a-fluidic CD4 cell count is achieved by eliminating operational fluid flow via "moving the substrate", as opposed to "flowing liquid" in traditional ELISA or microfluidic methods. This is the first demonstration of capturing and detecting cells from unprocessed whole blood using the enzyme-linked immunosorbent assay (ELISA) in a microfluidic channel. Combined with cell phone imaging, the presented micro-a-fluidic ELISA platform holds great promise for offering rapid CD4 cell count to scale up much needed ART in resource-constrained settings. The developed system can be extended to multiple areas for ELISA-related assays.

  View details for DOI 10.1038/srep03796

  View details for Web of Science ID 000330044300001

  View details for PubMedID 24448112

  View details for PubMedCentralID PMC3898414

• Guided and magnetic self-assembly of tunable magnetoceptive gels. Nature communications Tasoglu, S., Yu, C. H., Gungordu, H. I., Guven, S., Vural, T., Demirci, U. 2014; 5: 4702-?

  Abstract

  Self-assembly of components into complex functional patterns at microscale is common in nature, and used increasingly in numerous disciplines such as optoelectronics, microfabrication, sensors, tissue engineering and computation. Here, we describe the use of stable radicals to guide the self-assembly of magnetically tunable gels, which we call 'magnetoceptive' materials at the scale of hundreds of microns to a millimeter, each can be programmed by shape and composition, into heterogeneous complex structures. Using paramagnetism of free radicals as a driving mechanism, complex heterogeneous structures are built in the magnetic field generated by permanent magnets. The overall magnetic signature of final structure is erased via an antioxidant vitamin E, subsequent to guided self-assembly. We demonstrate unique capabilities of radicals and antioxidants in fabrication of soft systems with heterogeneity in material properties, such as porosity, elastic modulus and mass density; then in bottom-up tissue engineering and finally, levitational and selective assembly of microcomponents.

  View details for DOI 10.1038/ncomms5702

  View details for PubMedID 25175148

• Untethered micro-robotic coding of three-dimensional material composition NATURE COMMUNICATIONS Tasoglu, S., Diller, E., Guven, S., Sitti, M., Demirci, U. 2014; 5

  Abstract

  Complex functional materials with three-dimensional micro- or nano-scale dynamic compositional features are prevalent in nature. However, the generation of three-dimensional functional materials composed of both soft and rigid microstructures, each programmed by shape and composition, is still an unsolved challenge. Here we describe a method to code complex materials in three-dimensions with tunable structural, morphological and chemical features using an untethered magnetic micro-robot remotely controlled by magnetic fields. This strategy allows the micro-robot to be introduced to arbitrary microfluidic environments for remote two- and three-dimensional manipulation. We demonstrate the coding of soft hydrogels, rigid copper bars, polystyrene beads and silicon chiplets into three-dimensional heterogeneous structures. We also use coded microstructures for bottom-up tissue engineering by generating cell-encapsulating constructs.

  View details for DOI 10.1038/ncomms4124

  View details for Web of Science ID 000331084400033

  View details for PubMedID 24469115

• Exhaustion of Racing Sperm in Nature-Mimicking Microfluidic Channels During Sorting SMALL Tasoglu, S., Safaee, H., Zhang, X., Kingsley, J. L., Catalano, P. N., Gurkan, U. A., Nureddin, A., Kayaalp, E., Anchan, R. M., Maas, R. L., Tuezel, E., Demirci, U. 2013; 9 (20): 3374-3384

  Abstract

  Fertilization is central to the survival and propagation of a species, however, the precise mechanisms that regulate the sperm's journey to the egg are not well understood. In nature, the sperm has to swim through the cervical mucus, akin to a microfluidic channel. Inspired by this, a simple, cost-effective microfluidic channel is designed on the same scale. The experimental results are supported by a computational model incorporating the exhaustion time of sperm.

  View details for DOI 10.1002/smll.201300020

  View details for Web of Science ID 000326017700003

  View details for PubMedID 23677651

• Acute On-Chip HIV Detection Through Label-Free Electrical Sensing of Viral Nano-Lysate SMALL Shafiee, H., Jahangir, M., Inci, F., Wang, S., Willenbrecht, R. B., Giguel, F. F., Tsibris, A. M., Kuritzkes, D. R., Demirci, U. 2013; 9 (15): 2553-2563

  Abstract

  Development of portable biosensors has broad applications in environmental monitoring, clinical diagnosis, public health, and homeland security. There is an unmet need for pathogen detection at the point-of-care (POC) using a fast, sensitive, inexpensive, and easy-to-use method that does not require complex infrastructure and well-trained technicians. For instance, detection of Human Immunodeficiency Virus (HIV-1) at acute infection stage has been challenging, since current antibody-based POC technologies are not effective due to low concentration of antibodies. In this study, we demonstrated for the first time a label-free electrical sensing method that can detect lysed viruses, i.e. viral nano-lysate, through impedance analysis, offering an alternative technology to the antibody-based methods such as dipsticks and Enzyme-linked Immunosorbent Assay (ELISA). The presented method is a broadly applicable platform technology that can potentially be adapted to detect multiple pathogens utilizing impedance spectroscopy for other infectious diseases including herpes, influenza, hepatitis, pox, malaria, and tuberculosis. The presented method offers a rapid and portable tool that can be used as a detection technology at the POC in resource-constrained settings, as well as hospital and primary care settings.

  View details for DOI 10.1002/smll.201202195

  View details for Web of Science ID 000327792600013

  View details for PubMedID 23447456

• Nanostructured substrates for isolation of circulating tumor cells NANO TODAY Wang, L., Asghar, W., Demirci, U., Wan, Y. 2013; 8 (4): 374-387

  View details for DOI 10.1016/j.nantod.2013.07.001

  View details for Web of Science ID 000324783300008

• Point-of-care assays for tuberculosis: Role of nanotechnology/microfluidics BIOTECHNOLOGY ADVANCES Wang, S., Inci, F., De Libero, G., Singhal, A., Demirci, U. 2013; 31 (4): 438-449

  Abstract

  Tuberculosis (TB) remains one of the most devastating infectious diseases and its eradication is still unattainable given the limitations of current technologies for diagnosis, treatment and prevention. The World Health Organization's goal to eliminate TB globally by 2050 remains an ongoing challenge as delayed diagnosis and misdiagnosis of TB continue to fuel the worldwide epidemic. Despite considerable improvements in diagnostics for the last few decades, a simple and effective point-of-care TB diagnostic test is yet not available. Here, we review the current assays used for TB diagnosis, and highlight the recent advances in nanotechnology and microfluidics that potentially enable new approaches for TB diagnosis in resource-constrained settings.

  View details for DOI 10.1016/j.biotechadv.2013.01.006

  View details for Web of Science ID 000317453600003

  View details for PubMedID 23357365

• Nanoplasmonic Quantitative Detection of Intact Viruses from Unprocessed Whole Blood ACS NANO Inci, F., Tokel, O., Wang, S., Gurkan, U. A., Tasoglu, S., Kuritzkes, D. R., Demirci, U. 2013; 7 (6): 4733-4745

  Abstract

  Infectious diseases such as HIV and hepatitis B pose an omnipresent threat to global health. Reliable, fast, accurate, and sensitive platforms that can be deployed at the point-of-care (POC) in multiple settings, such as airports and offices, for detection of infectious pathogens are essential for the management of epidemics and possible biological attacks. To the best of our knowledge, no viral load technology adaptable to the POC settings exists today due to critical technical and biological challenges. Here, we present for the first time a broadly applicable technology for quantitative, nanoplasmonic-based intact virus detection at clinically relevant concentrations. The sensing platform is based on unique nanoplasmonic properties of nanoparticles utilizing immobilized antibodies to selectively capture rapidly evolving viral subtypes. We demonstrate the capture, detection, and quantification of multiple HIV subtypes (A, B, C, D, E, G, and subtype panel) with high repeatability, sensitivity, and specificity down to 98 ± 39 copies/mL (i.e., HIV subtype D) using spiked whole blood samples and clinical discarded HIV-infected patient whole blood samples validated by the gold standard, i.e., RT-qPCR. This platform technology offers an assay time of 1 h and 10 min (1 h for capture, 10 min for detection and data analysis). The presented platform is also able to capture intact viruses at high efficiency using immuno-surface chemistry approaches directly from whole blood samples without any sample preprocessing steps such as spin-down or sorting. Evidence is presented showing the system to be accurate, repeatable, and reliable. Additionally, the presented platform technology can be broadly adapted to detect other pathogens having reasonably well-described biomarkers by adapting the surface chemistry. Thus, this broadly applicable detection platform holds great promise to be implemented at POC settings, hospitals, and primary care settings.

  View details for DOI 10.1021/nn3036232

  View details for Web of Science ID 000321093800006

  View details for PubMedID 23688050

  View details for PubMedCentralID PMC3700402

• Flow induces epithelial-mesenchymal transition, cellular heterogeneity and biomarker modulation in 3D ovarian cancer nodules PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Rizvi, I., Gurkan, U. A., Tasoglu, S., Alagic, N., Celli, J. P., Mensah, L. B., Mai, Z., Demirci, U., Hasan, T. 2013; 110 (22): E1974-E1983

  Abstract

  Seventy-five percent of patients with epithelial ovarian cancer present with advanced-stage disease that is extensively disseminated intraperitoneally and prognosticates the poorest outcomes. Primarily metastatic within the abdominal cavity, ovarian carcinomas initially spread to adjacent organs by direct extension and then disseminate via the transcoelomic route to distant sites. Natural fluidic streams of malignant ascites triggered by physiological factors, including gravity and negative subdiaphragmatic pressure, carry metastatic cells throughout the peritoneum. We investigated the role of fluidic forces as modulators of metastatic cancer biology in a customizable microfluidic platform using 3D ovarian cancer nodules. Changes in the morphological, genetic, and protein profiles of biomarkers associated with aggressive disease were evaluated in the 3D cultures grown under controlled and continuous laminar flow. A modulation of biomarker expression and tumor morphology consistent with increased epithelial-mesenchymal transition, a critical step in metastatic progression and an indicator of aggressive disease, is observed because of hydrodynamic forces. The increase in epithelial-mesenchymal transition is driven in part by a posttranslational up-regulation of epidermal growth factor receptor (EGFR) expression and activation, which is associated with the worst prognosis in ovarian cancer. A flow-induced, transcriptionally regulated decrease in E-cadherin protein expression and a simultaneous increase in vimentin is observed, indicating increased metastatic potential. These findings demonstrate that fluidic streams induce a motile and aggressive tumor phenotype. The microfluidic platform developed here potentially provides a flow-informed framework complementary to conventional mechanism-based therapeutic strategies, with broad applicability to other lethal malignancies.

  View details for DOI 10.1073/pnas.1216989110

  View details for Web of Science ID 000320500000003

  View details for PubMedID 23645635

• Paramagnetic Levitational Assembly of Hydrogels ADVANCED MATERIALS Tasoglu, S., Kavaz, D., Gurkan, U. A., Guven, S., Chen, P., Zheng, R., Demirci, U. 2013; 25 (8): 1137-1143

  View details for DOI 10.1002/adma.201200285

  View details for Web of Science ID 000315102600007

  View details for PubMedID 23288557

• Simple Precision Creation of Digitally Specified, Spatially Heterogeneous, Engineered Tissue Architectures ADVANCED MATERIALS Gurkan, U. A., Fan, Y., Xu, F., Erkmen, B., Urkac, E. S., Parlakgul, G., Bernstein, J., Xing, W., Boyden, E. S., Demirci, U. 2013; 25 (8): 1192-1198

  View details for DOI 10.1002/adma.201203261

  View details for Web of Science ID 000315102600017

  View details for PubMedID 23192949

• Bioprinting for stem cell research TRENDS IN BIOTECHNOLOGY Tasoglu, S., Demirci, U. 2013; 31 (1): 10-19

  Abstract

  Recently, there has been growing interest in applying bioprinting techniques to stem cell research. Several bioprinting methods have been developed utilizing acoustics, piezoelectricity, and lasers to deposit living cells onto receiving substrates. Using these technologies, spatially defined gradients of immobilized biomolecules can be engineered to direct stem cell differentiation into multiple subpopulations of different lineages. Stem cells can also be patterned in a high-throughput manner onto flexible implementation patches for tissue regeneration or onto substrates with the goal of accessing encapsulated stem cells of interest for genomic analysis. Here, we review recent achievements with bioprinting technologies in stem cell research, and identify future challenges and potential applications including tissue engineering and regenerative medicine, wound healing, and genomics.

  View details for DOI 10.1016/j.tibtech.2012.10.005

  View details for Web of Science ID 000314014600005

  View details for PubMedID 23260439

• Manipulating biological agents and cells in micro-scale volumes for applications in medicine CHEMICAL SOCIETY REVIEWS Tasoglu, S., Gurkan, U. A., Wang, S., Demirci, U. 2013; 42 (13): 5788-5808

  Abstract

  Recent technological advances provide new tools to manipulate cells and biological agents in micro/nano-liter volumes. With precise control over small volumes, the cell microenvironment and other biological agents can be bioengineered; interactions between cells and external stimuli can be monitored; and the fundamental mechanisms such as cancer metastasis and stem cell differentiation can be elucidated. Technological advances based on the principles of electrical, magnetic, chemical, optical, acoustic, and mechanical forces lead to novel applications in point-of-care diagnostics, regenerative medicine, in vitro drug testing, cryopreservation, and cell isolation/purification. In this review, we first focus on the underlying mechanisms of emerging examples for cell manipulation in small volumes targeting applications such as tissue engineering. Then, we illustrate how these mechanisms impact the aforementioned biomedical applications, discuss the associated challenges, and provide perspectives for further development.

  View details for DOI 10.1039/c3cs60042d

  View details for Web of Science ID 000320120000009

  View details for PubMedID 23575660

• Prediction and control of number of cells in microdroplets by stochastic modeling LAB ON A CHIP Ceyhan, E., Xu, F., Gurkan, U. A., Emre, A. E., Turali, E. S., El Assal, R., Acikgenc, A., Wu, C. M., Demirci, U. 2012; 12 (22): 4884-4893

  Abstract

  Manipulation and encapsulation of cells in microdroplets has found many applications in various fields such as clinical diagnostics, pharmaceutical research, and regenerative medicine. The control over the number of cells in individual droplets is important especially for microfluidic and bioprinting applications. There is a growing need for modeling approaches that enable control over a number of cells within individual droplets. In this study, we developed statistical models based on negative binomial regression to determine the dependence of number of cells per droplet on three main factors: cell concentration in the ejection fluid, droplet size, and cell size. These models were based on experimental data obtained by using a microdroplet generator, where the presented statistical models estimated the number of cells encapsulated in droplets. We also propose a stochastic model for the total volume of cells per droplet. The statistical and stochastic models introduced in this study are adaptable to various cell types and cell encapsulation technologies such as microfluidic and acoustic methods that require reliable control over number of cells per droplet provided that settings and interaction of the variables is similar.

  View details for DOI 10.1039/c2lc40523g

  View details for Web of Science ID 000310865200039

  View details for PubMedID 23034772

• Smart Interface Materials Integrated with Microfluidics for On-Demand Local Capture and Release of Cells ADVANCED HEALTHCARE MATERIALS Gurkan, U. A., Tasoglu, S., Akkaynak, D., Avci, O., Unluisler, S., Canikyan, S., Maccallum, N., Demirci, U. 2012; 1 (5): 661-668

  Abstract

  Stimuli responsive, smart interface materials are integrated with microfluidic technologies creating new functions for a broad range of biological and clinical applications by controlling the material and cell interactions. Local capture and on-demand local release of cells are demonstrated with spatial and temporal control in a microfluidic system.

  View details for DOI 10.1002/adhm.201200009

  View details for Web of Science ID 000315114500016

  View details for PubMedID 23184803

• Bioprinting anisotropic stem cell microenvironment Gurkan, U. A., Sung, Y., El Assal, R., Xu, F., Trachtenberg, A., Kuo, W., Demirci, U. WILEY-BLACKWELL. 2012: 366–366

  View details for Web of Science ID 000308313003155

• Release of Magnetic Nanoparticles from Cell-Encapsulating Biodegradable Nanobiomaterials ACS NANO Xu, F., Inci, F., Mullick, O., Gurkan, U. A., Sung, Y., Kavaz, D., Li, B., Denkbas, E. B., Demirci, U. 2012; 6 (8): 6640-6649

  Abstract

  The future of tissue engineering requires development of intelligent biomaterials using nanoparticles. Magnetic nanoparticles (MNPs) have several applications in biology and medicine; one example is Food and Drug Administration (FDA)-approved contrast agents in magnetic resonance imaging. Recently, MNPs have been encapsulated within cell-encapsulating hydrogels to create novel nanobiomaterials (i.e., M-gels), which can be manipulated and assembled in magnetic fields. The M-gels can be used as building blocks for bottom-up tissue engineering to create 3D tissue constructs. For tissue engineering applications of M-gels, it is essential to study the release of encapsulated MNPs from the hydrogel polymer network and the effect of MNPs on hydrogel properties, including mechanical characteristics, porosity, swelling behavior, and cellular response (e.g., viability, growth). Therefore, we evaluated the release of MNPs from photocrosslinkable gelatin methacrylate hydrogels as the polymer network undergoes biodegradation using inductively coupled plasma atomic emission spectroscopy. MNP release correlated linearly with hydrogel biodegradation rate with correlation factors (Pearson product moment correlation coefficient) of 0.96 ± 0.03 and 0.99 ± 0.01 for MNP concentrations of 1% and 5%, respectively. We also evaluated the effect of MNPs on hydrogel mechanical properties, porosity, and swelling behavior, as well as cell viability and growth in MNP-encapsulating hydrogels. Fibroblasts encapsulated with MNPs in hydrogels remained viable (>80% at t = 144 h) and formed microtissue constructs in culture (t = 144 h). These results indicated that MNP-encapsulating hydrogels show promise as intelligent nanobiomaterials, with great potential to impact broad areas of bioengineering, including tissue engineering, regenerative medicine, and pharmaceutical applications.

  View details for DOI 10.1021/nn300902w

  View details for Web of Science ID 000307988900015

  View details for PubMedID 22680777

• Nanoliter droplet vitrification for oocyte cryopreservation NANOMEDICINE Zhang, X., Khimji, I., Shao, L., Safaee, H., Desai, K., Keles, H. O., Gurkan, U. A., Kayaalp, E., Nurreddin, A., Anchan, R. M., Maas, R. L., Demirci, U. 2012; 7 (4): 553-564

  Abstract

  Oocyte cryopreservation remains largely experimental, with live birth rates of only 2-4% per thawed oocyte. In this study, we present a nanoliter droplet technology for oocyte vitrification.An ejector-based droplet vitrification system was designed to continuously cryopreserve oocytes in nanoliter droplets. Oocyte survival rates, morphologies and parthenogenetic development after each vitrification step were assessed in comparison with fresh oocytes.Oocytes were retrieved after cryoprotectant agent loading/unloading, and nanoliter droplet encapsulation showed comparable survival rates to fresh oocytes after 24 h in culture. Also, oocytes recovered after vitrification/thawing showed similar morphologies to those of fresh oocytes. Additionally, the rate of oocyte parthenogenetic activation after nanoliter droplet encapsulation was comparable with that observed for fresh oocytes. This nanoliter droplet technology enables the vitrification of oocytes at higher cooling and warming rates using lower cryoprotectant agent levels (i.e., 1.4 M ethylene glycol, 1.1 M dimethyl sulfoxide and 1 M sucrose), thus making it a potential technology to improve oocyte cryopreservation outcomes.

  View details for DOI 10.2217/NNM.11.145

  View details for Web of Science ID 000303076000012

  View details for PubMedID 22188180

• Emerging Technologies for Assembly of Microscale Hydrogels ADVANCED HEALTHCARE MATERIALS Gurkan, U. A., Tasoglu, S., Kavaz, D., Demirel, M. C., Demirci, U. 2012; 1 (2): 149-158

  Abstract

  Assembly of cell encapsulating building blocks (i.e., microscale hydrogels) has significant applications in areas including regenerative medicine, tissue engineering, and cell-based in vitro assays for pharmaceutical research and drug discovery. Inspired by the repeating functional units observed in native tissues and biological systems (e.g., the lobule in liver, the nephron in kidney), assembly technologies aim to generate complex tissue structures by organizing microscale building blocks. Novel assembly technologies enable fabrication of engineered tissue constructs with controlled properties including tunable microarchitectural and predefined compositional features. Recent advances in micro- and nano-scale technologies have enabled engineering of microgel based three dimensional (3D) constructs. There is a need for high-throughput and scalable methods to assemble microscale units with a complex 3D micro-architecture. Emerging assembly methods include novel technologies based on microfluidics, acoustic and magnetic fields, nanotextured surfaces, and surface tension. In this review, we survey emerging microscale hydrogel assembly methods offering rapid, scalable microgel assembly in 3D, and provide future perspectives and discuss potential applications.

  View details for DOI 10.1002/adhm.201200011

  View details for Web of Science ID 000315111100002

  View details for PubMedID 23184717

• Efficient on-chip isolation of HIV subtypes LAB ON A CHIP Wang, S., Esfahani, M., Gurkan, U. A., Inci, F., Kuritzkes, D. R., Demirci, U. 2012; 12 (8): 1508-1515

  Abstract

  HIV has caused a global pandemic over the last three decades. There is an unmet need to develop point-of-care (POC) viral load diagnostics to initiate and monitor antiretroviral treatment in resource-constrained settings. Particularly, geographical distribution of HIV subtypes poses significant challenges for POC immunoassays. Here, we demonstrated a microfluidic device that can effectively capture various subtypes of HIV particles through anti-gp120 antibodies, which were immobilized on the microchannel surface. We first optimized an antibody immobilization process using fluorescent antibodies, quantum dot staining and AFM studies. The results showed that anti-gp120 antibodies were immobilized on the microchannel surface with an elevated antibody density and uniform antibody orientation using a Protein G-based surface chemistry. Further, RT-qPCR analysis showed that HIV particles of subtypes A, B and C were captured repeatably with high efficiencies of 77.2 ± 13.2%, 82.1 ± 18.8, and 80.9 ± 14.0% from culture supernatant, and 73.2 ± 13.6, 74.4 ± 14.6 and 78.3 ± 13.3% from spiked whole blood at a viral load of 1000 copies per mL, respectively. HIV particles of subtypes A, B and C were captured with high efficiencies of 81.8 ± 9.4%, 72.5 ± 18.7, and 87.8 ± 3.2% from culture supernatant, and 74.6 ± 12.9, 75.5 ± 6.7 and 69.7 ± 9.5% from spiked whole blood at a viral load of 10,000 copies per mL, respectively. The presented immuno-sensing device enables the development of POC on-chip technologies to monitor viral load and guide antiretroviral treatment (ART) in resource-constrained settings.

  View details for DOI 10.1039/c2lc20706k

  View details for Web of Science ID 000301986500016

  View details for PubMedID 22391989

• Sheathless Size-Based Acoustic Particle Separation SENSORS Guldiken, R., Jo, M. C., Gallant, N. D., Demirci, U., Zhe, J. 2012; 12 (1): 905-922

  Abstract

  Particle separation is of great interest in many biological and biomedical applications. Flow-based methods have been used to sort particles and cells. However, the main challenge with flow based particle separation systems is the need for a sheath flow for successful operation. Existence of the sheath liquid dilutes the analyte, necessitates precise flow control between sample and sheath flow, requires a complicated design to create sheath flow and separation efficiency depends on the sheath liquid composition. In this paper, we present a microfluidic platform for sheathless particle separation using standing surface acoustic waves. In this platform, particles are first lined up at the center of the channel without introducing any external sheath flow. The particles are then entered into the second stage where particles are driven towards the off-center pressure nodes for size based separation. The larger particles are exposed to more lateral displacement in the channel due to the acoustic force differences. Consequently, different-size particles are separated into multiple collection outlets. The prominent feature of the present microfluidic platform is that the device does not require the use of the sheath flow for positioning and aligning of particles. Instead, the sheathless flow focusing and separation are integrated within a single microfluidic device and accomplished simultaneously. In this paper, we demonstrated two different particle size-resolution separations; (1) 3 μm and 10 μm and (2) 3 μm and 5 μm. Also, the effects of the input power, the flow rate, and particle concentration on the separation efficiency were investigated. These technologies have potential to impact broadly various areas including the essential microfluidic components for lab-on-a-chip system and integrated biological and biomedical applications.

  View details for DOI 10.3390/s120100905

  View details for Web of Science ID 000299537100047

  View details for PubMedID 22368502

• Portable microfluidic chip for detection of Escherichia coli in produce and blood INTERNATIONAL JOURNAL OF NANOMEDICINE Wang, S., Inci, F., Chaunzwa, T. L., Ramanujam, A., Vasudevan, A., Subramanian, S., Ip, A. C., Sridharan, B., Gurkan, U. A., Demirci, U. 2012; 7: 2591-2600

  Abstract

  Pathogenic agents can lead to severe clinical outcomes such as food poisoning, infection of open wounds, particularly in burn injuries and sepsis. Rapid detection of these pathogens can monitor these infections in a timely manner improving clinical outcomes. Conventional bacterial detection methods, such as agar plate culture or polymerase chain reaction, are time-consuming and dependent on complex and expensive instruments, which are not suitable for point-of-care (POC) settings. Therefore, there is an unmet need to develop a simple, rapid method for detection of pathogens such as Escherichia coli. Here, we present an immunobased microchip technology that can rapidly detect and quantify bacterial presence in various sources including physiologically relevant buffer solution (phosphate buffered saline [PBS]), blood, milk, and spinach. The microchip showed reliable capture of E. coli in PBS with an efficiency of 71.8% ± 5% at concentrations ranging from 50 to 4,000 CFUs/mL via lipopolysaccharide binding protein. The limits of detection of the microchip for PBS, blood, milk, and spinach samples were 50, 50, 50, and 500 CFUs/mL, respectively. The presented technology can be broadly applied to other pathogens at the POC, enabling various applications including surveillance of food supply and monitoring of bacteriology in patients with burn wounds.

  View details for DOI 10.2147/IJN.S29629

  View details for Web of Science ID 000304615700001

  View details for PubMedID 22679370

  View details for PubMedCentralID PMC3368510

• Simple filter microchip for rapid separation of plasma and viruses from whole blood INTERNATIONAL JOURNAL OF NANOMEDICINE Wang, S., Sarenac, D., Chen, M. H., Huang, S., Giguel, F. F., Kuritzkes, D. R., Demirci, U. 2012; 7: 5019-5028

  Abstract

  Sample preparation is a significant challenge for detection and sensing technologies, since the presence of blood cells can interfere with the accuracy and reliability of virus detection at the nanoscale for point-of-care testing. To the best of our knowledge, there is not an existing on-chip virus isolation technology that does not use complex fluidic pumps. Here, we presented a lab-on-a-chip filter device to isolate plasma and viruses from unprocessed whole blood based on size exclusion without using a micropump. We demonstrated that viruses (eg, HIV) can be separated on a filter-based chip (2-μm pore size) from HIV-spiked whole blood at high recovery efficiencies of 89.9% ± 5.0%, 80.5% ± 4.3%, and 78.2% ± 3.8%, for viral loads of 1000, 10,000 and 100,000 copies/mL, respectively. Meanwhile, 81.7% ± 6.7% of red blood cells and 89.5% ± 2.4% of white blood cells were retained on 2 μm pore-sized filter microchips. We also tested these filter microchips with seven HIV-infected patient samples and observed recovery efficiencies ranging from 73.1% ± 8.3% to 82.5% ± 4.1%. These results are first steps towards developing disposable point-of-care diagnostics and monitoring devices for resource-constrained settings, as well as hospital and primary care settings.

  View details for DOI 10.2147/IJN.S32579

  View details for Web of Science ID 000308793900001

  View details for PubMedID 23055720

  View details for PubMedCentralID PMC3457680

• The assembly of cell-encapsulating microscale hydrogels using acoustic waves BIOMATERIALS Xu, F., Finley, T. D., Turkaydin, M., Sung, Y., Gurkan, U. A., Yavuz, A. S., Guldiken, R. O., Demirci, U. 2011; 32 (31): 7847-7855

  Abstract

  Microscale hydrogels find widespread applications in medicine and biology, e.g., as building blocks for tissue engineering and regenerative medicine. In these applications, these microgels are assembled to fabricate large complex 3D constructs. The success of this approach requires non-destructive and high throughput assembly of the microgels. Although various assembly methods have been developed based on modifying interfaces, and using microfluidics, so far, none of the available assembly technologies have shown the ability to assemble microgels using non-invasive fields rapidly within seconds in an efficient way. Acoustics has been widely used in biomedical arena to manipulate droplets, cells and biomolecules. In this study, we developed a simple, non-invasive acoustic assembler for cell-encapsulating microgels with maintained cell viability (>93%). We assessed the assembler for both microbeads (with diameter of 50 μm and 100 μm) and microgels of different sizes and shapes (e.g., cubes, lock-and-key shapes, tetris, saw) in microdroplets (with volume of 10 μL, 20 μL, 40 μL, 80 μL). The microgels were assembled in seconds in a non-invasive manner. These results indicate that the developed acoustic approach could become an enabling biotechnology tool for tissue engineering, regenerative medicine, pharmacology studies and high throughput screening applications.

  View details for DOI 10.1016/j.biomaterials.2011.07.010

  View details for Web of Science ID 000295072600011

  View details for PubMedID 21820734

• Three-Dimensional Magnetic Assembly of Microscale Hydrogels ADVANCED MATERIALS Xu, F., Wu, C. M., Rengarajan, V., Finley, T. D., Keles, H. O., Sung, Y., Li, B., Gurkan, U. A., Demirci, U. 2011; 23 (37): 4254-4260

  View details for DOI 10.1002/adma.201101962

  View details for Web of Science ID 000296258900005

  View details for PubMedID 21830240

• Microengineering methods for cell-based microarrays and high-throughput drug-screening applications BIOFABRICATION Xu, F., Wu, J., Wang, S., Durmus, N. G., Gurkan, U. A., Demirci, U. 2011; 3 (3)

  Abstract

  Screening for effective therapeutic agents from millions of drug candidates is costly, time consuming, and often faces concerns due to the extensive use of animals. To improve cost effectiveness, and to minimize animal testing in pharmaceutical research, in vitro monolayer cell microarrays with multiwell plate assays have been developed. Integration of cell microarrays with microfluidic systems has facilitated automated and controlled component loading, significantly reducing the consumption of the candidate compounds and the target cells. Even though these methods significantly increased the throughput compared to conventional in vitro testing systems and in vivo animal models, the cost associated with these platforms remains prohibitively high. Besides, there is a need for three-dimensional (3D) cell-based drug-screening models which can mimic the in vivo microenvironment and the functionality of the native tissues. Here, we present the state-of-the-art microengineering approaches that can be used to develop 3D cell-based drug-screening assays. We highlight the 3D in vitro cell culture systems with live cell-based arrays, microfluidic cell culture systems, and their application to high-throughput drug screening. We conclude that among the emerging microengineering approaches, bioprinting holds great potential to provide repeatable 3D cell-based constructs with high temporal, spatial control and versatility.

  View details for DOI 10.1088/1758-5082/3/3/034101

  View details for Web of Science ID 000294955200003

  View details for PubMedID 21725152

• Transport of a soft cargo on a nanoscale ratchet APPLIED PHYSICS LETTERS Sekeroglu, K., Gurkan, U. A., Demirci, U., Demirel, M. C. 2011; 99 (6)

  Abstract

  Surface ratchets can guide droplet transport for microfluidic systems. Here, we demonstrated the actuation of microgels encapsulated in droplets using a unidirectional nanotextured surface, which moves droplets with low vibration amplitudes by a ratcheting mechanism. The nanofilm carries droplets along the ratchets with minimal drop shape deformation to move the encapsulated soft cargo, i.e., microscale hydrogels. The tilted nanorods of the nanofilm produce unidirectional wetting, thereby enabling droplet motion in a single direction. Maximum droplet translation speed on the nanofilm was determined to be 3.5 mm∕s, which offers a pathway towards high throughput microgel assembly applications to build complex constructs.

  View details for DOI 10.1063/1.3625430

  View details for Web of Science ID 000293857700092

  View details for PubMedID 21901051

• Emerging technologies in medical applications of minimum volume vitrification NANOMEDICINE Zhang, X., Catalano, P. N., Gurkan, U. A., Khimji, I., Demirci, U. 2011; 6 (6): 1115-1129

  Abstract

  Cell/tissue biopreservation has broad public health and socio-economic impact affecting millions of lives. Cryopreservation technologies provide an efficient way to preserve cells and tissues targeting the clinic for applications including reproductive medicine and organ transplantation. Among these technologies, vitrification has displayed significant improvement in post-thaw cell viability and function by eliminating harmful effects of ice crystal formation compared to the traditional slow freezing methods. However, high cryoprotectant agent concentrations are required, which induces toxicity and osmotic stress to cells and tissues. It has been shown that vitrification using small sample volumes (i.e., <1 µl) significantly increases cooling rates and hence reduces the required cryoprotectant agent levels. Recently, emerging nano- and micro-scale technologies have shown potential to manipulate picoliter to nanoliter sample sizes. Therefore, the synergistic integration of nanoscale technologies with cryogenics has the potential to improve biopreservation methods.

  View details for DOI 10.2217/NNM.11.71

  View details for Web of Science ID 000295697600020

  View details for PubMedID 21955080

• Statistical Modeling of Single Target Cell Encapsulation PLOS ONE Moon, S., Ceyhan, E., Gurkan, U. A., Demirci, U. 2011; 6 (7)

  Abstract

  High throughput drop-on-demand systems for separation and encapsulation of individual target cells from heterogeneous mixtures of multiple cell types is an emerging method in biotechnology that has broad applications in tissue engineering and regenerative medicine, genomics, and cryobiology. However, cell encapsulation in droplets is a random process that is hard to control. Statistical models can provide an understanding of the underlying processes and estimation of the relevant parameters, and enable reliable and repeatable control over the encapsulation of cells in droplets during the isolation process with high confidence level. We have modeled and experimentally verified a microdroplet-based cell encapsulation process for various combinations of cell loading and target cell concentrations. Here, we explain theoretically and validate experimentally a model to isolate and pattern single target cells from heterogeneous mixtures without using complex peripheral systems.

  View details for DOI 10.1371/journal.pone.0021580

  View details for Web of Science ID 000292956800005

  View details for PubMedID 21814548

• Enumeration of CD4(+) T-Cells Using a Portable Microchip Count Platform in Tanzanian HIV-Infected Patients PLOS ONE Moon, S., Gurkan, U. A., Blander, J., Fawzi, W. W., Aboud, S., Mugusi, F., Kuritzkes, D. R., Demirci, U. 2011; 6 (7)

  Abstract

  CD4(+) T-lymphocyte count (CD4 count) is a standard method used to monitor HIV-infected patients during anti-retroviral therapy (ART). The World Health Organization (WHO) has pointed out or recommended that a handheld, point-of-care, reliable, and affordable CD4 count platform is urgently needed in resource-scarce settings.HIV-infected patient blood samples were tested at the point-of-care using a portable and label-free microchip CD4 count platform that we have developed. A total of 130 HIV-infected patient samples were collected that included 16 de-identified left over blood samples from Brigham and Women's Hospital (BWH), and 114 left over samples from Muhimbili University of Health and Allied Sciences (MUHAS) enrolled in the HIV and AIDS care and treatment centers in the City of Dar es Salaam, Tanzania. The two data groups from BWH and MUHAS were analyzed and compared to the commonly accepted CD4 count reference method (FACSCalibur system).The portable, battery operated and microscope-free microchip platform developed in our laboratory (BWH) showed significant correlation in CD4 counts compared with FACSCalibur system both at BWH (r = 0.94, p<0.01) and MUHAS (r = 0.49, p<0.01), which was supported by the Bland-Altman methods comparison analysis. The device rapidly produced CD4 count within 10 minutes using an in-house developed automated cell counting program.We obtained CD4 counts of HIV-infected patients using a portable platform which is an inexpensive (<$1 material cost) and disposable microchip that uses whole blood sample (<10 µl) without any pre-processing. The system operates without the need for antibody-based fluorescent labeling and expensive fluorescent illumination and microscope setup. This portable CD4 count platform displays agreement with the FACSCalibur results and has the potential to expand access to HIV and AIDS monitoring using fingerprick volume of whole blood and helping people who suffer from HIV and AIDS in resource-limited settings.

  View details for DOI 10.1371/journal.pone.0021409

  View details for Web of Science ID 000292632000012

  View details for PubMedID 21754988

• Embryonic stem cell bioprinting for uniform and controlled size embryoid body formation BIOMICROFLUIDICS Xu, F., Sridharan, B., Wang, S., Gurkan, U. A., Syverud, B., Demirci, U. 2011; 5 (2)

  Abstract

  Embryonic stem cells (ESCs) are pluripotent with multilineage potential to differentiate into virtually all cell types in the organism and thus hold a great promise for cell therapy and regenerative medicine. In vitro differentiation of ESCs starts with a phase known as embryoid body (EB) formation. EB mimics the early stages of embryogenesis and plays an essential role in ESC differentiation in vitro. EB uniformity and size are critical parameters that directly influence the phenotype expression of ESCs. Various methods have been developed to form EBs, which involve natural aggregation of cells. However, challenges persist to form EBs with controlled size, shape, and uniformity in a reproducible manner. The current hanging-drop methods are labor intensive and time consuming. In this study, we report an approach to form controllable, uniform-sized EBs by integrating bioprinting technologies with the existing hanging-drop method. The approach presented here is simple, robust, and rapid. We present significantly enhanced EB size uniformity compared to the conventional manual hanging-drop method.

  View details for DOI 10.1063/1.3580752

  View details for Web of Science ID 000292329700012

  View details for PubMedID 21799713

• Automated and Adaptable Quantification of Cellular Alignment from Microscopic Images for Tissue Engineering Applications TISSUE ENGINEERING PART C-METHODS Xu, F., Beyazoglu, T., Hefner, E., Gurkan, U. A., Demirci, U. 2011; 17 (6): 641-649

  Abstract

  Cellular alignment plays a critical role in functional, physical, and biological characteristics of many tissue types, such as muscle, tendon, nerve, and cornea. Current efforts toward regeneration of these tissues include replicating the cellular microenvironment by developing biomaterials that facilitate cellular alignment. To assess the functional effectiveness of the engineered microenvironments, one essential criterion is quantification of cellular alignment. Therefore, there is a need for rapid, accurate, and adaptable methodologies to quantify cellular alignment for tissue engineering applications. To address this need, we developed an automated method, binarization-based extraction of alignment score (BEAS), to determine cell orientation distribution in a wide variety of microscopic images. This method combines a sequenced application of median and band-pass filters, locally adaptive thresholding approaches and image processing techniques. Cellular alignment score is obtained by applying a robust scoring algorithm to the orientation distribution. We validated the BEAS method by comparing the results with the existing approaches reported in literature (i.e., manual, radial fast Fourier transform-radial sum, and gradient based approaches). Validation results indicated that the BEAS method resulted in statistically comparable alignment scores with the manual method (coefficient of determination R(2)=0.92). Therefore, the BEAS method introduced in this study could enable accurate, convenient, and adaptable evaluation of engineered tissue constructs and biomaterials in terms of cellular alignment and organization.

  View details for DOI 10.1089/ten.tec.2011.0038

  View details for Web of Science ID 000291202200003

  View details for PubMedID 21370940

• Living Bacterial Sacrificial Porogens to Engineer Decellularized Porous Scaffolds PLOS ONE Xu, F., Sridharan, B., Durmus, N. G., Wang, S., Yavuz, A. S., Gurkan, U. A., Demirci, U. 2011; 6 (4)

  Abstract

  Decellularization and cellularization of organs have emerged as disruptive methods in tissue engineering and regenerative medicine. Porous hydrogel scaffolds have widespread applications in tissue engineering, regenerative medicine and drug discovery as viable tissue mimics. However, the existing hydrogel fabrication techniques suffer from limited control over pore interconnectivity, density and size, which leads to inefficient nutrient and oxygen transport to cells embedded in the scaffolds. Here, we demonstrated an innovative approach to develop a new platform for tissue engineered constructs using live bacteria as sacrificial porogens. E.coli were patterned and cultured in an interconnected three-dimensional (3D) hydrogel network. The growing bacteria created interconnected micropores and microchannels. Then, the scafold was decellularized, and bacteria were eliminated from the scaffold through lysing and washing steps. This 3D porous network method combined with bioprinting has the potential to be broadly applicable and compatible with tissue specific applications allowing seeding of stem cells and other cell types.

  View details for DOI 10.1371/journal.pone.0019344

  View details for Web of Science ID 000290020700050

  View details for PubMedID 21552485

• Drop-on-Demand Single Cell Isolation and Total RNA Analysis PLOS ONE Moon, S., Kim, Y., Dong, L., Lombardi, M., Haeggstrom, E., Jensen, R. V., Hsiao, L., Demirci, U. 2011; 6 (3)

  Abstract

  Technologies that rapidly isolate viable single cells from heterogeneous solutions have significantly contributed to the field of medical genomics. Challenges remain both to enable efficient extraction, isolation and patterning of single cells from heterogeneous solutions as well as to keep them alive during the process due to a limited degree of control over single cell manipulation. Here, we present a microdroplet based method to isolate and pattern single cells from heterogeneous cell suspensions (10% target cell mixture), preserve viability of the extracted cells (97.0±0.8%), and obtain genomic information from isolated cells compared to the non-patterned controls. The cell encapsulation process is both experimentally and theoretically analyzed. Using the isolated cells, we identified 11 stem cell markers among 1000 genes and compare to the controls. This automated platform enabling high-throughput cell manipulation for subsequent genomic analysis employs fewer handling steps compared to existing methods.

  View details for DOI 10.1371/journal.pone.0017455

  View details for Web of Science ID 000288247800007

  View details for PubMedID 21412416

• Blood Banking in Living Droplets PLOS ONE Samot, J., Moon, S., Shao, L., Zhang, X., Xu, F., Song, Y., Keles, H. O., Matloff, L., Markel, J., Demirci, U. 2011; 6 (3)

  Abstract

  Blood banking has a broad public health impact influencing millions of lives daily. It could potentially benefit from emerging biopreservation technologies. However, although vitrification has shown advantages over traditional cryopreservation techniques, it has not been incorporated into transfusion medicine mainly due to throughput challenges. Here, we present a scalable method that can vitrify red blood cells in microdroplets. This approach enables the vitrification of large volumes of blood in a short amount of time, and makes it a viable and scalable biotechnology tool for blood cryopreservation.

  View details for DOI 10.1371/journal.pone.0017530

  View details for Web of Science ID 000288247800010

  View details for PubMedID 21412411

• A three-dimensional in vitro ovarian cancer coculture model using a high-throughput cell patterning platform BIOTECHNOLOGY JOURNAL Xu, F., Celli, J., Rizvi, I., Moon, S., Hasan, T., Demirci, U. 2011; 6 (2): 204-212

  Abstract

  In vitro 3D cancer models that provide a more accurate representation of disease in vivo are urgently needed to improve our understanding of cancer pathology and to develop better cancer therapies. However, development of 3D models that are based on manual ejection of cells from micropipettes suffer from inherent limitations such as poor control over cell density, limited repeatability, low throughput, and, in the case of coculture models, lack of reproducible control over spatial distance between cell types (e.g., cancer and stromal cells). In this study, we build on a recently introduced 3D model in which human ovarian cancer (OVCAR-5) cells overlaid on Matrigel™ spontaneously form multicellular acini. We introduce a high-throughput automated cell printing system to bioprint a 3D coculture model using cancer cells and normal fi broblasts micropatterned on Matrigel™ . Two cell types were patterned within a spatially controlled microenvironment (e.g., cell density, cell-cell distance) in a high-throughput and reproducible manner; both cell types remained viable during printing and continued to proliferate following patterning. This approach enables the miniaturization of an established macro-scale 3D culture model and would allow systematic investigation into the multiple unknown regulatory feedback mechanisms between tumor and stromal cells and provide a tool for high-throughput drug screening.

  View details for DOI 10.1002/biot.201000340

  View details for Web of Science ID 000287718500009

  View details for PubMedID 21298805

• Miniaturized lensless imaging systems for cell and microorganism visualization in point-of-care testing BIOTECHNOLOGY JOURNAL Gurkan, U. A., Moon, S., Geckil, H., Xu, F., Wang, S., Lu, T. J., Demirci, U. 2011; 6 (2): 138-149

  Abstract

  Low-cost, robust, and user-friendly diagnostic capabilities at the point-of-care (POC) are critical for treating infectious diseases and preventing their spread in developing countries. Recent advances in micro- and nanoscale technologies have enabled the merger of optical and fluidic technologies (optofluidics) paving the way for cost-effective lensless imaging and diagnosis for POC testing in resource-limited settings. Applications of the emerging lensless imaging technologies include detecting and counting cells of interest, which allows rapid and affordable diagnostic decisions. This review presents the advances in lensless imaging and diagnostic systems, and their potential clinical applications in developing countries. The emerging technologies are reviewed from a POC perspective considering cost effectiveness, portability, sensitivity, throughput and ease of use for resource-limited settings.

  View details for DOI 10.1002/biot.201000427

  View details for Web of Science ID 000287718500002

  View details for PubMedID 21298800

• Integration of cell phone imaging with microchip ELISA to detect ovarian cancer HE4 biomarker in urine at the point-of-care LAB ON A CHIP Wang, S., Zhao, X., Khimji, I., Akbas, R., Qiu, W., Edwards, D., Cramer, D. W., Ye, B., Demirci, U. 2011; 11 (20): 3411-3418

  Abstract

  Ovarian cancer is asymptomatic in the early stages and most patients present with advanced levels of disease. The lack of cost-effective methods that can achieve frequent, simple and non-invasive testing hinders early detection and causes high mortality in ovarian cancer patients. Here, we report a simple and inexpensive microchip ELISA-based detection module that employs a portable detection system, i.e., a cell phone/charge-coupled device (CCD) to quantify an ovarian cancer biomarker, HE4, in urine. Integration of a mobile application with a cell phone enabled immediate processing of microchip ELISA results, which eliminated the need for a bulky, expensive spectrophotometer. The HE4 level detected by a cell phone or a lensless CCD system was significantly elevated in urine samples from cancer patients (n = 19) than healthy controls (n = 20) (p < 0.001). Receiver operating characteristic (ROC) analyses showed that the microchip ELISA coupled with a cell phone running an automated analysis mobile application had a sensitivity of 89.5% at a specificity of 90%. Under the same specificity, the microchip ELISA coupled with a CCD had a sensitivity of 84.2%. In conclusion, integration of microchip ELISA with cell phone/CCD-based colorimetric measurement technology can be used to detect HE4 biomarker at the point-of-care (POC), paving the way to create bedside technologies for diagnostics and treatment monitoring.

  View details for DOI 10.1039/c1lc20479c

  View details for Web of Science ID 000295270100007

  View details for PubMedID 21881677

• Lensless imaging for simultaneous microfluidic sperm monitoring and sorting LAB ON A CHIP Zhang, X., Khimji, I., Gurkan, U. A., Safaee, H., Catalano, P. N., Keles, H. O., Kayaalp, E., Demirci, U. 2011; 11 (15): 2535-2540

  Abstract

  5.3 million American couples of reproductive age (9%) are affected by infertility, among which male factors account for up to 50% of cases, which necessitates the identification of parameters defining sperm quality, including sperm count and motility. In vitro fertilization (IVF) with or without intra cytoplasmic sperm injection (ICSI) has become the most widely used assisted reproductive technology (ART) in modern clinical practice to overcome male infertility challenges. One of the obstacles of IVF and ICSI lies in identifying and isolating the most motile and presumably healthiest sperm from semen samples that have low sperm counts (oligozoospermia) and/or low sperm motility (oligospermaesthenia). Microfluidic systems have shown potential to sort sperm with flow systems. However, the small field of view (FOV) of conventional microscopes commonly used to image sperm motion presents challenges in tracking a large number of sperm cells simultaneously. To address this challenge, we have integrated a lensless charge-coupled device (CCD) with a microfluidic chip to enable wide FOV and automatic recording as the sperm move inside a microfluidic channel. The integrated system enables the sorting and tracking of a population of sperm that have been placed in a microfluidic channel. This channel can be monitored in both horizontal and vertical configuration similar to a swim-up column method used clinically. Sperm motilities can be quantified by tracing the shadow paths for individual sperm. Moreover, as the sperm are sorted by swimming from the inlet towards the outlet of a microfluidic channel, motile sperm that reach the outlet can be extracted from the channel at the end of the process. This technology can lead to methods to evaluate each sperm individually in terms of motility response in a wide field of view, which could prove especially useful, when working with oligozoospermic or oligospermaesthenic samples, in which the most motile sperm need to be isolated from a pool of small number of sperm.

  View details for DOI 10.1039/c1lc20236g

  View details for Web of Science ID 000292966700009

  View details for PubMedID 21677993

• Controlled viable release of selectively captured label-free cells in microchannels LAB ON A CHIP Gurkan, U. A., Anand, T., Tas, H., Elkan, D., Akay, A., Keles, H. O., Demirci, U. 2011; 11 (23): 3979-3989

  Abstract

  Selective capture of cells from bodily fluids in microchannels has broadly transformed medicine enabling circulating tumor cell isolation, rapid CD4(+) cell counting for HIV monitoring, and diagnosis of infectious diseases. Although cell capture methods have been demonstrated in microfluidic systems, the release of captured cells remains a significant challenge. Viable retrieval of captured label-free cells in microchannels will enable a new era in biological sciences by allowing cultivation and post-processing. The significant challenge in release comes from the fact that the cells adhere strongly to the microchannel surface, especially when immuno-based immobilization methods are used. Even though fluid shear and enzymes have been used to detach captured cells in microchannels, these methods are known to harm cells and affect cellular characteristics. This paper describes a new technology to release the selectively captured label-free cells in microchannels without the use of fluid shear or enzymes. We have successfully released the captured CD4(+) cells (3.6% of the mononuclear blood cells) from blood in microfluidic channels with high specificity (89% ± 8%), viability (94% ± 4%), and release efficiency (59% ± 4%). We have further validated our system by specifically capturing and controllably releasing the CD34(+) stem cells from whole blood, which were quantified to be 19 cells per million blood cells in the blood samples used in this study. Our results also indicated that both CD4(+) and CD34(+) cells released from the microchannels were healthy and amenable for in vitro culture. Manual flow based microfluidic method utilizes inexpensive, easy to fabricate microchannels allowing selective label-free cell capture and release in less than 10 minutes, which can also be used at the point-of-care. The presented technology can be used to isolate and purify a broad spectrum of cells from mixed populations offering widespread applications in applied biological sciences, such as tissue engineering, regenerative medicine, rare cell and stem cell isolation, proteomic/genomic research, and clonal/population analyses.

  View details for DOI 10.1039/c1lc20487d

  View details for Web of Science ID 000296737100007

  View details for PubMedID 22002065

• Advances in developing HIV-1 viral load assays for resource-limited settings BIOTECHNOLOGY ADVANCES Wang, S., Xu, F., Demirci, U. 2010; 28 (6): 770-781

  Abstract

  Commercial HIV-1 RNA viral load assays have been routinely used in developed countries to monitor antiretroviral treatment (ART). However, these assays require expensive equipment and reagents, well-trained operators, and established laboratory infrastructure. These requirements restrict their use in resource-limited settings where people are most afflicted with the HIV-1 epidemic. Inexpensive alternatives such as the Ultrasensitive p24 assay, the reverse transcriptase (RT) assay and in-house reverse transcription quantitative polymerase chain reaction (RT-qPCR) have been developed. However, they are still time-consuming, technologically complex and inappropriate for decentralized laboratories as point-of-care (POC) tests. Recent advances in microfluidics and nanotechnology offer new strategies to develop low-cost, rapid, robust and simple HIV-1 viral load monitoring systems. We review state-of-the-art technologies used for HIV-1 viral load monitoring in both developed and developing settings. Emerging approaches based on microfluidics and nanotechnology, which have potential to be integrated into POC HIV-1 viral load assays, are also discussed.

  View details for DOI 10.1016/j.biotechadv.2010.06.004

  View details for Web of Science ID 000283526500012

  View details for PubMedID 20600784

• Impact of a compound droplet on a flat surface: A model for single cell epitaxy PHYSICS OF FLUIDS Tasoglu, S., Kaynak, G., Szeri, A. J., Demirci, U., Muradoglu, M. 2010; 22 (8)

  Abstract

  The impact and spreading of a compound viscous droplet on a flat surface are studied computationally using a front-tracking method as a model for the single cell epitaxy. This is a technology developed to create two-dimensional and three-dimensional tissue constructs cell by cell by printing cell-encapsulating droplets precisely on a substrate using an existing ink-jet printing method. The success of cell printing mainly depends on the cell viability during the printing process, which requires a deeper understanding of the impact dynamics of encapsulated cells onto a solid surface. The present study is a first step in developing a model for deposition of cell-encapsulating droplets. The inner droplet representing the cell, the encapsulating droplet, and the ambient fluid are all assumed to be Newtonian. Simulations are performed for a range of dimensionless parameters to probe the deformation and rate of deformation of the encapsulated cell, which are both hypothesized to be related to cell damage. The deformation of the inner droplet consistently increases: as the Reynolds number increases; as the diameter ratio of the encapsulating droplet to the cell decreases; as the ratio of surface tensions of the air-solution interface to the solution-cell interface increases; as the viscosity ratio of the cell to encapsulating droplet decreases; or as the equilibrium contact angle decreases. It is observed that maximum deformation for a range of Weber numbers has (at least) one local minimum at We=2. Thereafter, the effects of cell deformation on viability are estimated by employing a correlation based on the experimental data of compression of cells between parallel plates. These results provide insight into achieving optimal parameter ranges for maximal cell viability during cell printing.

  View details for DOI 10.1063/1.3475527

  View details for Web of Science ID 000281905900010

  View details for PubMedID 20838481

• Microporous Cell-Laden Hydrogels for Engineered Tissue Constructs BIOTECHNOLOGY AND BIOENGINEERING Park, J. H., Chung, B. G., Lee, W. G., Kim, J., Brigham, M. D., Shim, J., Lee, S., Hwang, C. M., Durmus, N. G., Demirci, U., Khademhosseini, A. 2010; 106 (1): 138-148

  Abstract

  In this article, we describe an approach to generate microporous cell-laden hydrogels for fabricating biomimetic tissue engineered constructs. Micropores at different length scales were fabricated in cell-laden hydrogels by micromolding fluidic channels and leaching sucrose crystals. Microengineered channels were created within cell-laden hydrogel precursors containing agarose solution mixed with sucrose crystals. The rapid cooling of the agarose solution was used to gel the solution and form micropores in place of the sucrose crystals. The sucrose leaching process generated homogeneously distributed micropores within the gels, while enabling the direct immobilization of cells within the gels. We also characterized the physical, mechanical, and biological properties (i.e., microporosity, diffusivity, and cell viability) of cell-laden agarose gels as a function of engineered porosity. The microporosity was controlled from 0% to 40% and the diffusivity of molecules in the porous agarose gels increased as compared to controls. Furthermore, the viability of human hepatic carcinoma cells that were cultured in microporous agarose gels corresponded to the diffusion profile generated away from the microchannels. Based on their enhanced diffusive properties, microporous cell-laden hydrogels containing a microengineered fluidic channel can be a useful tool for generating tissue structures for regenerative medicine and drug discovery applications.

  View details for DOI 10.1002/bit.22667

  View details for Web of Science ID 000276844500014

  View details for PubMedID 20091766

• Engineering hydrogels as extracellular matrix mimics NANOMEDICINE Geckil, H., Xu, F., Zhang, X., Moon, S., Demirci, U. 2010; 5 (3): 469-484

  Abstract

  Extracellular matrix (ECM) is a complex cellular environment consisting of proteins, proteoglycans, and other soluble molecules. ECM provides structural support to mammalian cells and a regulatory milieu with a variety of important cell functions, including assembling cells into various tissues and organs, regulating growth and cell-cell communication. Developing a tailored in vitro cell culture environment that mimics the intricate and organized nanoscale meshwork of native ECM is desirable. Recent studies have shown the potential of hydrogels to mimic native ECM. Such an engineered native-like ECM is more likely to provide cells with rational cues for diagnostic and therapeutic studies. The research for novel biomaterials has led to an extension of the scope and techniques used to fabricate biomimetic hydrogel scaffolds for tissue engineering and regenerative medicine applications. In this article, we detail the progress of the current state-of-the-art engineering methods to create cell-encapsulating hydrogel tissue constructs as well as their applications in in vitro models in biomedicine.

  View details for DOI 10.2217/NNM.10.12

  View details for Web of Science ID 000277074000015

  View details for PubMedID 20394538

• Nano/Microfluidics for diagnosis of infectious diseases in developing countries ADVANCED DRUG DELIVERY REVIEWS Lee, W. G., Kim, Y., Chung, B. G., Demirci, U., Khademhosseini, A. 2010; 62 (4-5): 449-457

  Abstract

  Nano/Microfluidic technologies are emerging as powerful enabling tools for diagnosis and monitoring of infectious diseases in both developed and developing countries. Miniaturized nano/microfluidic platforms that precisely manipulate small fluid volumes can be used to enable medical diagnosis in a more rapid and accurate manner. In particular, these nano/microfluidic diagnostic technologies are potentially applicable to global health applications, since they are disposable, inexpensive, portable, and easy-to-use for detection of infectious diseases. In this paper, we review recent advances in nano/microfluidic technologies for clinical point-of-care applications at resource-limited settings in developing countries.

  View details for DOI 10.1016/j.addr.2009.11.016

  View details for Web of Science ID 000276123400007

  View details for PubMedID 19954755

  View details for PubMedCentralID PMC2829381

• Vitrification and levitation of a liquid droplet on liquid nitrogen PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Song, Y. S., Adler, D., Xu, F., Kayaalp, E., Nureddin, A., Anchan, R. M., Maas, R. L., Demirci, U. 2010; 107 (10): 4596-4600

  Abstract

  The vitrification of a liquid occurs when ice crystal formation is prevented in the cryogenic environment through ultrarapid cooling. In general, vitrification entails a large temperature difference between the liquid and its surrounding medium. In our droplet vitrification experiments, we observed that such vitrification events are accompanied by a Leidenfrost phenomenon, which impedes the heat transfer to cool the liquid, when the liquid droplet comes into direct contact with liquid nitrogen. This is distinct from the more generally observed Leidenfrost phenomenon that occurs when a liquid droplet is self-vaporized on a hot plate. In the case of rapid cooling, the phase transition from liquid to vitrified solid (i.e., vitrification) and the levitation of droplets on liquid nitrogen (i.e., Leidenfrost phenomenon) take place simultaneously. Here, we investigate these two simultaneous physical events by using a theoretical model containing three dimensionless parameters (i.e., Stefan, Biot, and Fourier numbers). We explain theoretically and observe experimentally a threshold droplet radius during the vitrification of a cryoprotectant droplet in the presence of the Leidenfrost effect.

  View details for DOI 10.1073/pnas.0914059107

  View details for Web of Science ID 000275368400020

  View details for PubMedID 20176969

• A droplet-based building block approach for bladder smooth muscle cell (SMC) proliferation BIOFABRICATION Xu, F., Moon, S. J., Emre, A. E., Turali, E. S., Song, Y. S., Hacking, S. A., Nagatomi, J., Demirci, U. 2010; 2 (1)

  Abstract

  Tissue engineering based on building blocks is an emerging method to fabricate 3D tissue constructs. This method requires depositing and assembling building blocks (cell-laden microgels) at high throughput. The current technologies (e.g., molding and photolithography) to fabricate microgels have throughput challenges and provide limited control over building block properties (e.g., cell density). The cell-encapsulating droplet generation technique has potential to address these challenges. In this study, we monitored individual building blocks for viability, proliferation and cell density. The results showed that (i) SMCs can be encapsulated in collagen droplets with high viability (>94.2 +/- 3.2%) for four cases of initial number of cells per building block (i.e. 7 +/- 2, 16 +/- 2, 26 +/- 3 and 37 +/- 3 cells/building block). (ii) Encapsulated SMCs can proliferate in building blocks at rates that are consistent (1.49 +/- 0.29) across all four cases, compared to that of the controls. (iii) By assembling these building blocks, we created an SMC patch (5 mm x 5 mm x 20 microm), which was cultured for 51 days forming a 3D tissue-like construct. The histology of the cultured patch was compared to that of a native rat bladder. These results indicate the potential of creating 3D tissue models at high throughput in vitro using building blocks.

  View details for DOI 10.1088/1758-5082/2/1/014105

  View details for Web of Science ID 000278118400006

  View details for PubMedID 20811120

• Multi-scale heat and mass transfer modelling of cell and tissue cryopreservation PHILOSOPHICAL TRANSACTIONS OF THE ROYAL SOCIETY A-MATHEMATICAL PHYSICAL AND ENGINEERING SCIENCES Xu, F., Moon, S., Zhang, X., Shao, L., Song, Y. S., Demirci, U. 2010; 368 (1912): 561-583

  Abstract

  Cells and tissues undergo complex physical processes during cryopreservation. Understanding the underlying physical phenomena is critical to improve current cryopreservation methods and to develop new techniques. Here, we describe multi-scale approaches for modelling cell and tissue cryopreservation including heat transfer at macroscale level, crystallization, cell volume change and mass transport across cell membranes at microscale level. These multi-scale approaches allow us to study cell and tissue cryopreservation.

  View details for DOI 10.1098/rsta.2009.0248

  View details for Web of Science ID 000273233400003

  View details for PubMedID 20047939

• Layer by Layer Three-Dimensional Tissue Epitaxy by Cell-Laden Hydrogel Droplets TISSUE ENGINEERING PART C-METHODS Moon, S., Hasan, S. K., Song, Y. S., Xu, F., Keles, H. O., Manzur, F., Mikkilineni, S., Hong, J. W., Nagatomi, J., Haeggstrom, E., Khademhosseini, A., Demirci, U. 2010; 16 (1): 157-166

  Abstract

  The ability to bioengineer three-dimensional (3D) tissues is a potentially powerful approach to treat diverse diseases such as cancer, loss of tissue function, or organ failure. Traditional tissue engineering methods, however, face challenges in fabricating 3D tissue constructs that resemble the native tissue microvasculature and microarchitectures. We have developed a bioprinter that can be used to print 3D patches of smooth muscle cells (5 mm x 5 mm x 81 microm) encapsulated within collagen. Current inkjet printing systems suffer from loss of cell viability and clogging. To overcome these limitations, we developed a system that uses mechanical valves to print high viscosity hydrogel precursors containing cells. The bioprinting platform that we developed enables (i) printing of multilayered 3D cell-laden hydrogel structures (16.2 microm thick per layer) with controlled spatial resolution (proximal axis: 18.0 +/- 7.0 microm and distal axis: 0.5 +/- 4.9 microm), (ii) high-throughput droplet generation (1 s per layer, 160 droplets/s), (iii) cell seeding uniformity (26 +/- 2 cells/mm(2) at 1 million cells/mL, 122 +/- 20 cells/mm(2) at 5 million cells/mL, and 216 +/- 38 cells/mm(2) at 10 million cells/mL), and (iv) long-term viability in culture (>90%, 14 days). This platform to print 3D tissue constructs may be beneficial for regenerative medicine applications by enabling the fabrication of printed replacement tissues.

  View details for DOI 10.1089/ten.tec.2009.0179

  View details for Web of Science ID 000274125800016

  View details for PubMedID 19586367

• Quantum dot-based HIV capture and imaging in a microfluidic channel BIOSENSORS & BIOELECTRONICS Kim, Y., Moon, S., Kuritzkes, D. R., Demirci, U. 2009; 25 (1): 253-258

  Abstract

  Globally, over 33.2 million people who mostly live in developing countries with limited access to the appropriate medical care suffer from the human immunodeficiency virus (HIV) infection. We developed an on-chip HIV capture and imaging method using quantum dots (Qdots) from fingerprick volume (10 microl) of unprocessed HIV-infected patient whole blood in anti-gp120 antibody-immobilized microfluidic chip. Two-color Qdots (Qdot525 and Qdot655 streptavidin conjugates) were used to identify the captured HIV by simultaneous labeling the envelope gp120 glycoprotein and its high-mannose glycans. This dual-stain imaging technique using Qdots provides a new and effective tool for accurate identification of HIV particles from patient whole blood without any pre-processing. This on-chip HIV capture and imaging platform creates new avenues for point-of-care diagnostics and monitoring applications of infectious diseases.

  View details for DOI 10.1016/j.bios.2009.06.023

  View details for Web of Science ID 000270538400042

  View details for PubMedID 19665685

• Engineered 3D tissue models for cell-laden microfluidic channels ANALYTICAL AND BIOANALYTICAL CHEMISTRY Song, Y. S., Lin, R. L., Montesano, G., Durmus, N. G., Lee, G., Yoo, S., Kayaalp, E., Haeggstrom, E., Khademhosseini, A., Demirci, U. 2009; 395 (1): 185-193

  Abstract

  Delivery of nutrients and oxygen within three-dimensional (3D) tissue constructs is important to maintain cell viability. We built 3D cell-laden hydrogels to validate a new tissue perfusion model that takes into account nutrition consumption. The model system was analyzed by simulating theoretical nutrient diffusion into cell-laden hydrogels. We carried out a parametric study considering different microchannel sizes and inter-channel separation in the hydrogel. We hypothesized that nutrient consumption needs to be taken into account when optimizing the perfusion channel size and separation. We validated the hypothesis by experiments. We fabricated circular microchannels (r = 400 microm) in 3D cell-laden hydrogel constructs (R = 7.5 mm, volume = 5 ml). These channels were positioned either individually or in parallel within hydrogels to increase nutrient and oxygen transport as a way to improve cell viability. We quantified the spatial distribution of viable cells within 3D hydrogel scaffolds without channels and with single- and dual-perfusion microfluidic channels. We investigated quantitatively the cell viability as a function of radial distance from the channels using experimental data and mathematical modeling of diffusion profiles. Our simulations show that a large-channel radius as well as a large channel to channel distance diffuse nutrients farther through a 3D hydrogel. This is important since our results reveal that there is a close correlation between nutrient profiles and cell viability across the hydrogel.

  View details for DOI 10.1007/s00216-009-2935-1

  View details for Web of Science ID 000268866800021

  View details for PubMedID 19629459

• Integrating microfluidics and lensless imaging for point-of-care testing BIOSENSORS & BIOELECTRONICS Moon, S., Keles, H. O., Ozcan, A., Khademhosseini, A., Haeggstrom, E., Kuritzkes, D., Demirci, U. 2009; 24 (11): 3208-3214

  Abstract

  We demonstrate an integrated platform that merges a microfluidic chip with lensless imaging to target CD4(+) T-lymphocyte counts for HIV point-of-care testing at resource-limited settings. The chips were designed and fabricated simply with a laser cutter without using expensive cleanroom equipment. To capture CD4(+) T-lymphocytes from blood, anti-CD4 antibody was immobilized on only one side of the microfluidic chip. These captured cells were detected through an optically clear chip using a charge coupled device (CCD) sensor by lensless shadow imaging techniques. Gray scale image of the captured cells in a 24 mm x 4 mm x 50 microm microfluidic chip was obtained by the lensless imaging platform. The automatic cell counting software enumerated the captured cells in 3s. Captured cells were also imaged with a fluorescence microscope and manually counted to characterize functionality of the integrated platform. The integrated platform achieved 70.2+/-6.5% capture efficiency, 88.8+/-5.4% capture specificity for CD4(+) T-lymphocytes, 96+/-1.6% CCD efficiency, and 83.5+/-2.4% overall platform performance (n=9 devices) compared to the gold standard, i.e. flow cytometry count. The integrated system gives a CD4 count from blood within 10 min. The integrated platform points a promising direction for point-of-care testing (POCT) to rapidly capture, image and count subpopulations of cells from blood samples in an automated matter.

  View details for DOI 10.1016/j.bios.2009.03.037

  View details for Web of Science ID 000267577900005

  View details for PubMedID 19467854

• Microscale electroporation: challenges and perspectives for clinical applications INTEGRATIVE BIOLOGY Lee, W. G., Demirci, U., Khademhosseini, A. 2009; 1 (3): 242-251

  Abstract

  Microscale engineering plays a significant role in developing tools for biological applications by miniaturizing devices and providing controllable microenvironments for in vitro cell research. Miniaturized devices offer numerous benefits in comparison to their macroscale counterparts, such as lower use of expensive reagents, biomimetic environments, and the ability to manipulate single cells. Microscale electroporation is one of the main beneficiaries of microscale engineering as it provides spatial and temporal control of various electrical parameters. Microscale electroporation devices can be used to reduce limitations associated with the conventional electroporation approaches such as variations in the local pH, electric field distortion, sample contamination, and the difficulties in transfecting and maintaining the viability of desired cell types. Here, we present an overview of recent advances of the microscale electroporation methods and their applications in biology, as well as current challenges for its use for clinical applications. We categorize microscale electroporation into microchannel and microcapillary electroporation. Microchannel-based electroporation can be used for transfecting cells within microchannels under dynamic flow conditions in a controlled and high-throughput fashion. In contrast, microcapillary-based electroporation can be used for transfecting cells within controlled reaction chambers under static flow conditions. Using these categories we examine the use of microscale electroporation for clinical applications related to HIV-1, stem cells, cancer and other diseases and discuss the challenges in further advancing this technology for use in clinical medicine and biology.

  View details for DOI 10.1039/b819201d

  View details for Web of Science ID 000267113900002

  View details for PubMedID 20023735

• Cell Proliferation in Bioprinted Cell-Laden Collagen Droplets 35th Annual Northeast Bioengineering Conference Xu, F., Emre, A. E., Turali, E. S., Hasan, S. K., Moon, S., Nagatomi, J., Khademhosseini, A., Demirci, U. IEEE. 2009: 390–391

  View details for Web of Science ID 000268886300192

• Microfluidics for cryopreservation LAB ON A CHIP Song, Y. S., Moon, S., Hulli, L., Hasan, S. K., Kayaalp, E., Demirci, U. 2009; 9 (13): 1874-1881

  Abstract

  Minimizing cell damage throughout the cryopreservation process is critical to enhance the overall outcome. Osmotic shock sustained during the loading and unloading of cryoprotectants (CPAs) is a major source of cell damage during the cryopreservation process. We introduce a microfluidic approach to minimize osmotic shock to cells during cryopreservation. This approach allows us to control the loading and unloading of CPAs in microfluidic channels using diffusion and laminar flow. We provide a theoretical explanation of how the microfluidic approach minimizes osmotic shock in comparison to conventional cryopreservation protocols via cell membrane transport modeling. Finally, we show that biological experiments are consistent with the proposed mathematical model. The results indicate that our novel microfluidic-based approach improves post-thaw cell survivability by up to 25% on average over conventional cryopreservation protocols. The method developed in this study provides a platform to cryopreserve cells with higher viability, functionality, and minimal inter-technician variability. This method introduces microfluidic technologies to the field of biopreservation, opening the door to future advancements at the interface of these fields.

  View details for DOI 10.1039/b823062e

  View details for Web of Science ID 000267124400008

  View details for PubMedID 19532962

• Rapid automated cell quantification on HIV microfluidic devices LAB ON A CHIP Alyassin, M. A., Moon, S., Keles, H. O., Manzur, F., Lin, R. L., Haeggstrom, E., Kuritzkes, D. R., Demirci, U. 2009; 9 (23): 3364-3369

  Abstract

  Lab-chip device analysis often requires high throughput quantification of fluorescent cell images, obtained under different conditions of fluorescent intensity, illumination, focal depth, and optical magnification. Many laboratories still use manual counting--a tedious, expensive process prone to inter-observer variability. The manual counting process can be automated for fast and precise data gathering and reduced manual bias. We present a method to segment and count cells in microfluidic chips that are labeled with a single stain, or multiple stains, using image analysis techniques in Matlab and discuss its advantages over manual counting. Microfluidic based cell capturing devices for HIV monitoring were used to validate our method. Captured CD4(+) CD3(+) T lymphocytes were stained with DAPI, AF488-anti CD4, and AF647-anti CD3 for cell identification. Altogether 4788 (76 x 3 x 21) gray color images were obtained from devices using discarded 10 HIV infected patient whole blood samples (21 devices). We observed that the automatic method performs similarly to manual counting for a small number of cells. However, automated counting is more accurate and more than 100 times faster than manual counting for multiple-color stained cells, especially when large numbers of cells need to be quantified (>500 cells). The algorithm is fully automatic for subsequent microscope images that cover the full device area. It accounts for problems that generally occur in fluorescent lab-chip cell images such as: uneven background, overlapping cell images and cell detection with multiple stains. This method can be used in laboratories to save time and effort, and to increase cell counting accuracy of lab-chip devices for various applications, such as circulating tumor cell detection, cell detection in biosensors, and HIV monitoring devices, i.e. CD4 counts.

  View details for DOI 10.1039/b911882a

  View details for Web of Science ID 000271647400007

  View details for PubMedID 19904402

• Integrating Microfluidics and Lensless Imaging for Point-of-Care Testing 35th Annual Northeast Bioengineering Conference Moon, S., Keles, H. O., Khademhosseini, A., Kuritzkes, D., Demirci, U. IEEE. 2009: 32–33

  View details for Web of Science ID 000268886300013

• LAYER BY LAYER 3D TISSUE EPITAXY BY CELL LADEN HYDROGEL DROPLETS 35th Annual Northeast Bioengineering Conference Hasan, S. K., Moon, S., Song, Y. S., Keles, H. O., Anzur, F., Mikkilineni, S., Hong, J. W., Nagatomi, J., Haeggstrom, E., Khademhosseini, A., Demirci, U. IEEE. 2009: 366–367

  View details for Web of Science ID 000268886300180

• The effect of soluble surfactant on the transient motion of a buoyancy-driven bubble PHYSICS OF FLUIDS Tasoglu, S., Demirci, U., Muradoglu, M. 2008; 20 (4)

  View details for DOI 10.1063/1.2912441

  View details for Web of Science ID 000255456600015

• Ultra wide-field lens-free monitoring of cells on-chip LAB ON A CHIP Ozcan, A., Demirci, U. 2008; 8 (1): 98-106

  Abstract

  We experimentally and theoretically demonstrate the proof-of-principle of a new lens-free cell monitoring platform that involves using an opto-electronic sensor array to record the shadow image of cells onto the sensor plane. This technology can monitor/count cells over a field-of-view that is more than two orders of magnitude larger than that of a conventional light microscope. Furthermore, it does not require any mechanical scanning or optical elements, such as microscope objectives or lenses. We also show that this optical approach can conveniently be combined with microfluidic channels, enabling parallel on-chip monitoring of various different cell types, e.g., blood cells, NIH-3T3 fibroblasts, murine embryonic stem cells, AML-12 hepatocytes. An important application of this approach could be a miniaturized point-of-care technology to obtain CD4 T lymphocyte counts of HIV infected patients in resource limited settings.

  View details for DOI 10.1039/b713695a

  View details for Web of Science ID 000251771000021

  View details for PubMedID 18094767

• Microcirculation within grooved substrates regulates cell positioning and cell docking inside microfluidic channels LAB ON A CHIP Manbachi, A., Shrivastava, S., Cioffi, M., Chung, B. G., Moretti, M., Demirci, U., Yliperttula, M., Khademhosseini, A. 2008; 8 (5): 747-754

  Abstract

  Immobilization of cells inside microfluidic devices is a promising approach for enabling studies related to drug screening and cell biology. Despite extensive studies in using grooved substrates for immobilizing cells inside channels, a systematic study of the effects of various parameters that influence cell docking and retention within grooved substrates has not been performed. We demonstrate using computational simulations that the fluid dynamic environment within microgrooves significantly varies with groove width, generating microcirculation areas in smaller microgrooves. Wall shear stress simulation predicted that shear stresses were in the opposite direction in smaller grooves (25 and 50 microm wide) in comparison to those in wider grooves (75 and 100 microm wide). To validate the simulations, cells were seeded within microfluidic devices, where microgrooves of different widths were aligned perpendicularly to the direction of the flow. Experimental results showed that, as predicted, the inversion of the local direction of shear stress within the smaller grooves resulted in alignment of cells on two opposite sides of the grooves under the same flow conditions. Also, the amplitude of shear stress within microgrooved channels significantly influenced cell retainment in the channels. Therefore, our studies suggest that microscale shear stresses greatly influence cellular docking, immobilization, and retention in fluidic systems and should be considered for the design of cell-based microdevices.

  View details for DOI 10.1039/b718212k

  View details for Web of Science ID 000255276700016

  View details for PubMedID 18432345

• A microchip approach for practical label-free CD4+ T-cell counting of HIV-infected subjects in resource-poor settings JAIDS-JOURNAL OF ACQUIRED IMMUNE DEFICIENCY SYNDROMES Cheng, X., Irimia, D., Dixon, M., Ziperstein, J. C., Demirci, U., Zamir, L., Tompkins, R. G., Toner, M., Rodriguez, W. R. 2007; 45 (3): 257-261

  Abstract

  Simple affordable CD4 cell counting is urgently needed to stage and monitor HIV-infected patients in resource-limited settings. To address the limitations of current approaches, we designed a simple, label-free, and cost-effective CD4 cell counting device using microfluidic technology. We previously described the fabrication of a microfluidic system for high-efficiency isolation of pure populations of CD4+ T cells based on cell affinity chromatography operated under controlled flow. Here, we compare the performance of a microfluidic CD4 cell counting device against standard flow cytometry in 49 HIV-positive subjects over a wide range of absolute CD4 cell counts. We observed a close correlation between CD4 cell counts from the microchip device and measurements by flow cytometry, using unprocessed whole blood from HIV-positive adult subjects. Sensitivities for distinguishing clinically relevant thresholds of 200, 350, and 500 cells/microL are 0.86, 0.90, and 0.97, respectively. Specificity is 0.94 or higher at all thresholds. This device can serve as a functional cartridge for fast, accurate, affordable, and simple CD4 cell counting in resource-limited settings.

  View details for Web of Science ID 000247572900001

  View details for PubMedID 17414933

• A cell-laden microfluidic hydrogel LAB ON A CHIP Ling, Y., Rubin, J., Deng, Y., Huang, C., Demirci, U., Karp, J. M., Khademhosseini, A. 2007; 7 (6): 756-762

  Abstract

  The encapsulation of mammalian cells within the bulk material of microfluidic channels may be beneficial for applications ranging from tissue engineering to cell-based diagnostic assays. In this work, we present a technique for fabricating microfluidic channels from cell-laden agarose hydrogels. Using standard soft lithographic techniques, molten agarose was molded against a SU-8 patterned silicon wafer. To generate sealed and water-tight microfluidic channels, the surface of the molded agarose was heated at 71 degrees C for 3 s and sealed to another surface-heated slab of agarose. Channels of different dimensions were generated and it was shown that agarose, though highly porous, is a suitable material for performing microfluidics. Cells embedded within the microfluidic molds were well distributed and media pumped through the channels allowed the exchange of nutrients and waste products. While most cells were found to be viable upon initial device fabrication, only those cells near the microfluidic channels remained viable after 3 days, demonstrating the importance of a perfused network of microchannels for delivering nutrients and oxygen to maintain cell viability in large hydrogels. Further development of this technique may lead to the generation of biomimetic synthetic vasculature for tissue engineering, diagnostics, and drug screening applications.

  View details for DOI 10.1039/b615486g

  View details for Web of Science ID 000247566400024

  View details for PubMedID 17538718

• Cell detection and counting through cell lysate impedance spectroscopy in microfluidic devices LAB ON A CHIP Cheng, X., Liu, Y., Irimia, D., Demirci, U., Yang, L., Zamir, L., Rodriguez, W. R., Toner, M., Bashir, R. 2007; 7 (6): 746-755

  Abstract

  Cell-based microfluidic devices have attracted interest for a wide range of applications. While optical cell counting and flow cytometry-type devices have been reported extensively, sensitive and efficient non-optical methods to detect and quantify cells attached over large surface areas within microdevices are generally lacking. We describe an electrical method for counting cells based on the measurement of changes in conductivity of the surrounding medium due to ions released from surface-immobilized cells within a microfluidic channel. Immobilized cells are lysed using a low conductivity, hypotonic media and the resulting change in impedance is measured using surface patterned electrodes to detect and quantify the number of cells. We found that the bulk solution conductance increases linearly with the number of isolated cells contributing to solution ion concentration. The method of cell lysate impedance spectroscopy is sensitive enough to detect 20 cells microL(-1), and offers a simple and efficient method for detecting and enumerating cells within microfluidic devices for many applications including measurement of CD4 cell counts in HIV patients in resource-limited settings. To our knowledge, this is the most sensitive approach using non-optical setups to enumerate immobilized cells. The microfluidic device, capable of isolating specific cell types from a complex bio-fluidic and quantifying cell number, can serve as a single use cartridge for a hand-held instrument to provide simple, fast and affordable cell counting in point-of-care settings.

  View details for DOI 10.1039/b705082h

  View details for Web of Science ID 000247566400023

  View details for PubMedID 17538717

• Rewritable self-assembled long-period gratings in photonic bandgap fibers using microparticles OPTICS COMMUNICATIONS Ozcan, A., Demirci, U. 2007; 270 (2): 225-228

  View details for DOI 10.1016/j.optcom.2006.09.032

  View details for Web of Science ID 000244157700019

• A microfluidic device for practical label-free CD4+T cell counting of HIV-infected subjects LAB ON A CHIP Cheng, X., Irimia, D., Dixon, M., Sekine, K., Demirci, U., Zamir, L., Tompkins, R. G., Rodriguez, W., Toner, M. 2007; 7 (2): 170-178

  Abstract

  Practical HIV diagnostics are urgently needed in resource-limited settings. While HIV infection can be diagnosed using simple, rapid, lateral flow immunoassays, HIV disease staging and treatment monitoring require accurate counting of a particular white blood cell subset, the CD4(+) T lymphocyte. To address the limitations of current expensive, technically demanding and/or time-consuming approaches, we have developed a simple CD4 counting microfluidic device. This device uses cell affinity chromatography operated under differential shear flow to specifically isolate CD4(+) T lymphocytes with high efficiency directly from 10 microliters of unprocessed, unlabeled whole blood. CD4 counts are obtained under an optical microscope in a rapid, simple and label-free fashion. CD4 counts determined in our device matched measurements by conventional flow cytometry among HIV-positive subjects over a wide range of absolute CD4 counts (R(2) = 0.93). This CD4 counting microdevice can be used for simple, rapid and affordable CD4 counting in point-of-care and resource-limited settings.

  View details for DOI 10.1039/b612966h

  View details for Web of Science ID 000244616300003

  View details for PubMedID 17268618

• Cell encapsulating droplet vitrification LAB ON A CHIP Demirci, U., Montesano, G. 2007; 7 (11): 1428-1433

  Abstract

  The capability to encapsulate single cells in droplets while retaining high cell viability (>90%) has great impact on tissue engineering, high-throughput screening, as well as clinical diagnostics and therapeutics. We demonstrate a novel method to vitrify a small number of cells using cell-encapsulating droplets. The method allows vitrification at low cryoprotectant concentration (1.5 M propanediol and 0.5 M trehalose), similar to that used in slow freezing protocols. The method was successfully applied to five different mammalian cell types: AML-12 hepatocytes, NIH-3T3 fibroblasts, HL-1 cardiomyocytes, mouse embryonic stem cells, and RAJI cells.

  View details for DOI 10.1039/b705809h

  View details for Web of Science ID 000250428200013

  View details for PubMedID 17960267

• Single cell epitaxy by acoustic picolitre droplets LAB ON A CHIP Demirci, U., Montesano, G. 2007; 7 (9): 1139-1145

  Abstract

  The capability to encapsulate single to few cells with micrometre precision, high viability, and controlled directionality via a nozzleless ejection technology using a gentle acoustic field would have great impact on tissue engineering, high throughput screening, and clinical diagnostics. We demonstrate encapsulation of single cells (or a few cells) ejected from an open pool in acoustic picolitre droplets. We have developed this technology for the specific purpose of printing cells in various biological fluids, including PBS and agarose hydrogels used in tissue engineering. We ejected various cell types, including mouse embryonic stem cells, fibroblasts, AML-12 hepatocytes, human Raji cells, and HL-1 cardiomyocytes encapsulated in acoustic picolitre droplets of around 37 microm in diameter at rates varying from 1 to 10,000 droplets per second. At such high throughput levels, we demonstrated cell viabilities of over 89.8% across various cell types. Moreover, this ejection method is readily adaptable to other biological applications, such as extracting data from single cells and generating large cell populations from single cells. The technique described in the current study may also be applied to investigate stem cell differentiation at the single cell level, to direct tissue printing, and to isolating pure RNA or DNA from a single cell at the picolitre level. Overall, the techniques described have the potential for widespread impact on many high-throughput testing applications in the biological and health sciences.

  View details for DOI 10.1039/b704965j

  View details for Web of Science ID 000248917500007

  View details for PubMedID 17713612

• Acoustic picoliter droplets for emerging applications in semiconductor industry and biotechnology JOURNAL OF MICROELECTROMECHANICAL SYSTEMS Demirci, U. 2006; 15 (4): 957-966

  View details for DOI 10.1109/JMEMS.2006.878879

  View details for Web of Science ID 000239712600026

• Droplet-based photoresist deposition APPLIED PHYSICS LETTERS Demirci, U. 2006; 88 (14)

  View details for DOI 10.1063/1.2191087

  View details for Web of Science ID 000236612000121

• Direct etch method for microfludic channel and nanoheight post-fabrication by picoliter droplets APPLIED PHYSICS LETTERS Demirci, U., Toner, M. 2006; 88 (5)

  View details for DOI 10.1063/1.2170143

  View details for Web of Science ID 000235136700083

• Direct etch method for microfluidic channel and nano-height post fabrication by picoliter droplets MEMS 2006: 19TH IEEE INTERNATIONAL CONFERENCE ON MICRO ELECTRO MECHANICAL SYSTEMS, TECHNICAL DIGEST Demirci, U., Toner, M. 2006: 326-329

  View details for Web of Science ID 000236994500082

• Femtoliter to picoliter droplet generation for organic polymer deposition using single reservoir ejector arrays IEEE TRANSACTIONS ON SEMICONDUCTOR MANUFACTURING Demirci, U., Yaralioglu, G. G., Haeggstrom, E., Khuri-Yakub, B. T. 2005; 18 (4): 709-715

  View details for DOI 10.1109/TSM.2005.858500

  View details for Web of Science ID 000233379400031

• Picolitre acoustic droplet ejection by ferntosecond laser micromachined multiple-orifice membrane-based 2D ejector arrays ELECTRONICS LETTERS Demirci, U., Ozcan, A. 2005; 41 (22): 1219-1220

  View details for DOI 10.1049/el:20053208

  View details for Web of Science ID 000233454800013

• Picoliter droplets for spinless photoresist deposition REVIEW OF SCIENTIFIC INSTRUMENTS Demirci, U. 2005; 76 (6)

  View details for DOI 10.1063/1.1922867

  View details for Web of Science ID 000229962000094

• Coherent array imaging using phased subarmys. Part II: Simulations and experimental results IEEE TRANSACTIONS ON ULTRASONICS FERROELECTRICS AND FREQUENCY CONTROL Johnson, J. A., Oralkan, O., Ergun, S., Demirci, U., Karaman, M., Khuri-Yakub, B. T. 2005; 52 (1): 51-64

  Abstract

  The basic principles and theory of phased subarray (PSA) imaging imaging provides the flexibility of reducing the number of front-end hardware channels between that of classical synthetic aperture (CSA) imaging--which uses only one element per firing event--and full-phased array (FPA) imaging-which uses all elements for each firing. The performance of PSA generally ranges between that obtained by CSA and FPA using the same array, and depends on the amount of hardware complexity reduction. For the work described in this paper, we performed FPA, CSA, and PSA imaging of a resolution phantom using both simulated and experimental data from a 3-MHz, 3.2-cm, 128-element capacitive micromachined ultrasound transducer (CMUT) array. The simulated system point responses in the spatial and frequency domains are presented as a means of studying the effects of signal bandwidth, reconstruction filter size, and subsampling rate on the PSA system performance. The PSA and FPA sector-scanned images were reconstructed using the wideband experimental data with 80% fractional bandwidth, with seven 32-element subarrays used for PSA imaging. The measurements on the experimental sector images indicate that, at the transmit focal zone, the PSA method provides a 10% improvement in the 6-dB lateral resolution, and the axial point resolution of PSA imaging is identical to that of FPA imaging. The signal-to-noise ratio (SNR) of PSA image was 58.3 dB, 4.9 dB below that of the FPA image, and the contrast-to-noise ratio (CNR) is reduced by 10%. The simulated and experimental test results presented in this paper validate theoretical expectations and illustrate the flexibility of PSA imaging as a way to exchange SNR and frame rate for simplified front-end hardware.

  View details for Web of Science ID 000226812800007

  View details for PubMedID 15742562

• Acoustically actuated flextensional SixNy and single-crystal silicon 2-D micromachined ejector arrays IEEE TRANSACTIONS ON SEMICONDUCTOR MANUFACTURING Demirci, U., Yaralioglu, G. G., Haeggstrom, E., Percin, G., Ergun, S., Khuri-Yakub, B. T. 2004; 17 (4): 517-524

  View details for DOI 10.1109/tsm.2004.835714

  View details for Web of Science ID 000224995800007

• Forward-viewing CMUT arrays for medical Imaging IEEE TRANSACTIONS ON ULTRASONICS FERROELECTRICS AND FREQUENCY CONTROL Demirci, U., Ergun, A. S., Oralkan, O., Karaman, M., Khuri-Yakub, B. T. 2004; 51 (7): 887-895

  Abstract

  This paper reports the design and testing of forward-viewing annular arrays fabricated using capacitive micromachined ultrasonic transducer (CMUT) technology. Recent research studies have shown that CMUTs have broad frequency bandwidth and high-transduction efficiency. One- and two-dimensional CMUT arrays of various sizes already have been fabricated, and their viability for medical imaging applications has been demonstrated. We fabricated 64-element, forward-viewing annular arrays using the standard CMUT fabrication process and carried out experiments to measure the operating frequency, bandwidth, and transmit/receive efficiency of the array elements. The annular array elements, designed for imaging applications in the 20 MHz range, had a resonance frequency of 13.5 MHz in air. The immersion pulse-echo data collected from a plane reflector showed that the devices operate in the 5-26 MHz range with a fractional bandwidth of 135%. The output pressure at the surface of the transducer was measured to be 24 kPa/V. These values translate into a dynamic range of 131.5 dB for 1-V excitation in 1-Hz bandwidth with a commercial low noise receiving circuitry. The designed, forward-viewing annular CMUT array is suitable for mounting on the front surface of a cylindrical catheter probe and can provide Doppler information for measurement of blood flow and guiding information for navigation through blood vessels in intravascular ultrasound imaging.

  View details for Web of Science ID 000222678000018

  View details for PubMedID 15301009

• Phased subarray imaging for low-cost, wideband coherent array imaging IEEE International Ultrasonics Symposium Johnson, J. A., Oralkan, O., Ergun, A. S., Demirci, U., Karaman, M., Khuri-Yakub, B. T. IEEE. 2003: 1875–1878

  View details for Web of Science ID 000189492100435

• 2D acoustically actuated micromachined droplet ejector array IEEE International Ultrasonics Symposium Demirci, U., Yaralioglu, G. G., Haeggstrom, E., Percin, G., Khuri-Yakub, B. T. IEEE. 2003: 1983–1986

  View details for Web of Science ID 000189492100460

• Capacitive micromachined ultrasonic transducers: Next-generation arrays for acoustic imaging? IEEE TRANSACTIONS ON ULTRASONICS FERROELECTRICS AND FREQUENCY CONTROL Oralkan, O., Ergun, A. S., Johnson, J. A., Karaman, M., Demirci, U., Kaviani, K., Lee, T. H., Khuri-Yakub, B. T. 2002; 49 (11): 1596-1610

  Abstract

  Piezoelectric materials have dominated the ultrasonic transducer technology. Recently, capacitive micromachined ultrasonic transducers (CMUTs) have emerged as an alternative technology offering advantages such as wide bandwidth, ease of fabricating large arrays, and potential for integration with electronics. The aim of this paper is to demonstrate the viability of CMUTs for ultrasound imaging. We present the first pulse-echo phased array B-scan sector images using a 128-element, one-dimensional (1-D) linear CMUT array. We fabricated 64- and 128-element 1-D CMUT arrays with 100% yield and uniform element response across the arrays. These arrays have been operated in immersion with no failure or degradation in performance over the time. For imaging experiments, we built a resolution test phantom roughly mimicking the attenuation properties of soft tissue. We used a PC-based experimental system, including custom-designed electronic circuits to acquire the complete set of 128 x 128 RF A-scans from all transmit-receive element combinations. We obtained the pulse-echo frequency response by analyzing the echo signals from wire targets. These echo signals presented an 80% fractional bandwidth around 3 MHz, including the effect of attenuation in the propagating medium. We reconstructed the B-scan images with a sector angle of 90 degrees and an image depth of 210 mm through offline processing by using RF beamforming and synthetic phased array approaches. The measured 6-dB lateral and axial resolutions at 135 mm depth were 0.0144 radians and 0.3 mm, respectively. The electronic noise floor of the image was more than 50 dB below the maximum mainlobe magnitude. We also performed preliminary investigations on the effects of crosstalk among array elements on the image quality. In the near field, some artifacts were observable extending out from the array to a depth of 2 cm. A tail also was observed in the point spread function (PSF) in the axial direction, indicating the existence of crosstalk. The relative amplitude of this tail with respect to the mainlobe was less than -20 dB.

  View details for Web of Science ID 000179224100016

  View details for PubMedID 12484483

• Medical imaging using capacitive micromachined ultrasonic transducer arrays 1st Ultrasonics International Conference Johnson, J., Oralkan, O., Demirci, U., Ergun, S., Karaman, M., Khuri-Yakub, P. ELSEVIER SCIENCE BV. 2002: 471–76

  Abstract

  We are investigating the use of capacitive micromachined ultrasonic transducers (cMUT's) for use in medical imaging. We propose an ultrasound probe architecture designed to provide volumetric ultrasound imaging from within an endoscope channel. A complete automated experimental system has been implemented for testing the imaging performance of cMUT arrays. This PC-based system includes custom-designed circuit boards, a software interface, and resolution test phantoms. We have already fabricated 1D and 2D cMUT arrays, and tested the pulse-echo imaging characteristics of 1D arrays. Beamforming and image formation algorithms that aim to reduce the complexity of data acquisition hardware are tested via numerical simulations and using real data acquired from our system.

  View details for Web of Science ID 000176648000083

  View details for PubMedID 12159985

• Broadband capacitive micromachined ultrasonic transducers ranging from 10 kHz to 60 mHz for imaging arrays and more IEEE International Ultrasonic Symposium Ergun, A. S., Huang, Y., Cheng, C. H., Oralkan, O., Johnson, J., Jagannathan, H., Demirci, U., Yaralioglu, G. G., Karaman, M., Khuri-Yakub, B. T. IEEE. 2002: 1039–1043

  View details for Web of Science ID 000182111700232

• Fabrication and characterization of 1-dimensional and 2-dimensional capacitive micromachined ultrasonic transducer (CMUT) arrays for 2-dimensional and volumetric ultrasonic imaging MTS/IEEE Oceans 2002 Conference Ergun, A. S., Cheng, C. H., Demirci, U., Khuri-Yakub, B. T. IEEE. 2002: 2361–2367

  View details for Web of Science ID 000182293200371

• An ultrasonic volumetric scanner for image-guided surgery 15th International Congress and Exhibition on Computer Assisted Radiology and Surgery Johnson, J., Oralkan, O., Kaviani, K., Demirci, U., Karaman, M., Khuri-Yakub, P. ELSEVIER SCIENCE BV. 2001: 187–192

  View details for Web of Science ID 000173844800032

• Capacitive micromachined ultrasonic transducer arrays for medical imaging: Experimental results IEEE International Ultrasonic Symposium Demirci, U., Oralkan, O., Johnson, J. A., Ergun, A. S., Karaman, M., Khuri-Yakub, B. T. IEEE. 2001: 957–960

  View details for Web of Science ID 000176890800203