Dr. Demirci is currently a Professor at Stanford University School of Medicine with tenure at the Canary Center for Early Cancer Detection. Prior to his Stanford appointment, he was an Associate Professor of Medicine at Brigham and Women's Hospital, Harvard Medical School and at Harvard-MIT Division of Health Sciences and Technology serving at the Division of Biomedical Engineering, Division of Infectious Diseases and Renal Division. He leads a group of 20+ researchers focusing on micro- and nano-scale technologies. He received his B.S. degree in Electrical Engineering in 1999 as a James B. Angell Scholar (summa cum laude) from University of Michigan, Ann Arbor. He received his M.S. degree in 2001 in Electrical Engineering, M.S. degree in Management Science and Engineering in 2005, and Ph.D. in Electrical Engineering in 2005, all from Stanford University.
The Demirci Bio-Acoustic MEMS in Medicine Lab (BAMM) specializes in applying micro- and nanoscale technologies to problems in medicine at the interface between micro/nanoscale engineering and medicine. Our goal is apply innovative technologies to clinical problems. Our major research theme focuses on creating new microfluidic technology platforms targeting broad applications in medicine. In this interdisciplinary space at the convergence of engineering, biology and materials science, we create novel technologies for disposable point-of-care (POC) diagnostics and monitoring of infectious diseases, cancer and controlling cellular microenvironment in nanoliter droplets for biopreservation and microscale tissue engineering applications. These applications are unified around our expertise to test the limits of cell manipulation by establishing microfluidic platforms to provide solutions to real world problems at the clinic.
Our lab creates technologies to manipulate cells in nanoliter volumes to enable solutions for real world problems in medicine including applications in infectious disease diagnostics and monitoring for global health, cancer early detection, cell encapsulation in nanoliter droplets for cryobiology, and bottom-up tissue engineering. Dr. Demirci has published over 120 peer reviewed publications in journals including PNAS, Nature Communications, Advanced Materials, Small, Trends in Biotechnology, Chemical Society Reviews and Lab-chip, over 150 conference abstracts and proceedings, 10+ book chapters, and an edited book. His work was highlighted in Wired Magazine, Nature Photonics, Nature Medicine, MIT Technology Review, Reuters Health News, Science Daily, AIP News, BioTechniques, and Biophotonics. He is fellow-elect of the American Institute of Biological and Medical Engineering (AIMBE, 2017). His scientific work has been recognized by numerous national and international awards including the NSF Faculty Early Career Development (CAREER) Award (2012), the IEEE-EMBS Early Career Achievement Award (2012), Scientist of the year award from Stanford radiology Department (2017). He was selected as one of the world’s top 35 young innovators under the age of 35 (TR-35) by the MIT Technology Review at the age of 28. In 2004, he led a team that won the Stanford University Entrepreneur’s Challenge Competition and Global Start-up Competition in Singapore. His work has been translated to start-up companies including DxNow, KOEK Biotechnology and LEVITAS. There has been over 10,000 live births in the US, Europe and Turkey using the sperm selection technology that came out of Dr. Demirci's lab. He has been cited over 2500 times within the last two years (H index, 48).
Honors & Awards
Fellow-Elect, American Institute of Medical and Biomedical Engineers (AIMBE) (2016)
2nd Place Student and Investigator Section Oral Presentation, Tissue Engineering and Regenerative Medicine International Society (TERMIS)-Asia Pacific Meeting (2016)
StarTURK Award, Assembly of Turkish American Association (2014)
Bright Futures Award, Brigham and Women’s Hospital, Brigham Research Institute (2013)
Sharktank Competition, American Epilepsy Foundation (2013)
Faculty Early Career Development Award, NSF (2012)
Early Career Achievement Award, IEEE-EMBS (2012)
Partners in Excellence Award, Partners Health Care (2011)
Coulter Translational Research Award, Biomedical Engineering Society (BMES) (2011)
Engineering in Medicine and Biology Research Award for Translational Research, IEEE-Wyss Institute (2011)
Chinese Young Investigator Award, National Science Foundation of China (2010)
The Outstanding Young Persons of the World, Junior Chamber International (JCI) (2009)
Nano-Biotechnology Award, National Science Council of Turkey and The Turkish Industrialists’ and Businessmen’s Association (2007)
TR-35 Award-MIT, MIT Technology Review (2006)
Ministry of Education Award, Turkish Ministry of Education (2005)
Winner of Accenture Grand Prize, Singapore Business Plan Competition (2004)
1st Place, BASES Entrepreneur’s Challenge Business Plan Competition, Stanford University (2004)
Outstanding Paper Award, Transactions on Ultrasonic, Ferroelectrics, and Frequency Control, IEEE (2003)
Raymond William Barrow (RWB) Stephens Student Prize of Elsevier Science, Proceedings of Ultrasonic International (2001)
Phi Kappa Phi, National Honor Society, University of Michigan (1999)
Scholarship for Undergraduate Education, Turkish Ministry of Education (1996)
James B. Angell Scholar, University of Michigan (1999)
Boards, Advisory Committees, Professional Organizations
Co-founder and Scientific Advisor, DxNow Inc. (2013 - Present)
Co-founder and Scientific Advisor, LevitasBio (2017 - Present)
Co-founder and Scientific Advisor, Koek Biotech (2012 - Present)
Ph.D., Stanford University, Stanford, CA, Electrical Engineering (2005)
M.S., Stanford University, Stanford, CA, Management Science and Engineering (2005)
M.S., Stanford University, Stanford, CA, Electrical Engineering (2001)
B.S., University of Michigan, Ann Arbor, MI, Electrical Engineering (1999)
- Senior Capstone Design I
BIOE 141A (Aut)
Independent Studies (8)
- Directed Investigation
BIOE 392 (Aut, Win, Spr, Sum)
- Directed Reading in Radiology
RAD 299 (Win, Spr)
- Early Clinical Experience in Radiology
RAD 280 (Win, Spr)
- Graduate Research
RAD 399 (Aut, Win, Spr)
- Medical Scholars Research
RAD 370 (Win, Spr)
- Out-of-Department Graduate Research
BIO 300X (Aut, Win, Spr, Sum)
- Readings in Radiology Research
RAD 101 (Win, Spr)
- Undergraduate Research
RAD 199 (Win, Spr)
- Directed Investigation
3-D Microwell Array System for Culturing Virus Infected Tumor Cells
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
Dynamic Microenvironment Induces Phenotypic Plasticity of Esophageal Cancer Cells Under Flow
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
A high throughput approach for analysis of cell nuclear deformability at single cell level
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
Flexible Substrate-Based Devices for Point-of-Care Diagnostics.
Trends in biotechnology
2016; 34 (11): 909-921
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
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
2016; 103: 90-104
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
2016; 28 (18): 3543-?
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
2016; 16 (4): 449-459
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 2016; 8 (1)
Integrating Cell Phone Imaging with Magnetic Levitation (i-LEV) for Label-Free Blood Analysis at the Point-of-Living.
2016; 12 (9): 1222-1229
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
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.
2016; 31: 301-311
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 2016; 28 (1): 161-?
Toxicology Study of Single-walled Carbon Nanotubes and Reduced Graphene Oxide in Human Sperm.
2016; 6: 30270-?
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
- Engineering cancer microenvironments for in vitro 3-D tumor models MATERIALS TODAY 2015; 18 (10): 539-553
- Graphene-protein field effect biosensors: glucose sensing MATERIALS TODAY 2015; 18 (9): 513-522
- Deformation of a single mouse oocyte in a constricted microfluidic channel MICROFLUIDICS AND NANOFLUIDICS 2015; 19 (4): 883-890
- Portable lensless wide-field microscopy imaging platform based on digital inline holography and multi-frame pixel super-resolution LIGHT-SCIENCE & APPLICATIONS 2015; 4
Biotunable Acoustic Node Assembly of Organoids.
Advanced healthcare materials
2015; 4 (13): 1937-1943
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
2015; 112 (32): E4354-63
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
Hydrosoluble, UV-crosslinkable and injectable chitosan for patterned cell-laden microgel and rapid transdermal curing hydrogel in vivo
2015; 22: 59-69
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
2015; 4 (10): 1469-1476
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
2015; 112 (28): E3661-8
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
Levitational Image Cytometry with Temporal Resolution
2015; 27 (26): 3901-?
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).
2015; 27 (26): 3900-?
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
2015; 4 (10): 1420-?
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
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
Multiscale assembly for tissue engineering and regenerative medicine
TRENDS IN BIOTECHNOLOGY
2015; 33 (5): 269-279
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
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
Functional maintenance of differentiated embryoid bodies in microfluidic systems: a platform for personalized medicine.
Stem cells translational medicine
2015; 4 (3): 261-268
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
- Highlights from the latest articles in advanced biomanufacturing at micro- and nano-scale. Nanomedicine 2015; 10 (3): 347-350
- Advances in Nanotechnology and Microfluidics for Human Papillomavirus Diagnostics PROCEEDINGS OF THE IEEE 2015; 103 (2): 161-178
Emerging Technologies for Point-of-Care Management of HIV Infection
ANNUAL REVIEW OF MEDICINE, VOL 66
2015; 66: 387-405
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 2015; 14: 1778-1781
Paper and flexible substrates as materials for biosensing platforms to detect multiple biotargets.
2015; 5: 8719-?
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
Recent advances in micro/nanotechnologies for global control of hepatitis B infection
2015; 33 (1): 178-190
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.)
2015; 1256: 111-121
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
2014; 78: 105-117
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
- Two-dimensional numerical study of flow dynamics of a nucleated cell tethered under shear flow CHEMICAL ENGINEERING SCIENCE 2014; 119: 236-244
- Selection of Functional Human Sperm with Higher DNA Integrity and Fewer Reactive Oxygen Species ADVANCED HEALTHCARE MATERIALS 2014; 3 (10): 1671-1679
- Nanomechanical motion of Escherichia coli adhered to a surface APPLIED PHYSICS LETTERS 2014; 105 (11)
- Microscale Assembly Directed by Liquid-Based Template ADVANCED MATERIALS 2014; 26 (34): 5936-?
Bio-inspired cryo-ink preserves red blood cell phenotype and function during nanoliter vitrification.
2014; 26 (33): 5815-5822
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.
2014; 9 (7): 895-903
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
2014; 11 (7): 2151-2159
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
ELSEVIER SCIENCE INC. 2014: 2125–32
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 2014; 114 (11): 5728-5752
Nanostructured Optical Photonic Crystal Biosensor for HIV Viral Load Measurement
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
Micro-a-fluidics ELISA for Rapid CD4 Cell Count at the Point-of-Care
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
Guided and magnetic self-assembly of tunable magnetoceptive gels.
2014; 5: 4702-?
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
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
2013; 9 (20): 3374-3384
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
2013; 9 (15): 2553-2563
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 2013; 8 (4): 374-387
Point-of-care assays for tuberculosis: Role of nanotechnology/microfluidics
2013; 31 (4): 438-449
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
2013; 7 (6): 4733-4745
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
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
2013; 110 (22): E1974-E1983
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 2013; 25 (8): 1137-1143
- Simple Precision Creation of Digitally Specified, Spatially Heterogeneous, Engineered Tissue Architectures ADVANCED MATERIALS 2013; 25 (8): 1192-1198
Bioprinting for stem cell research
TRENDS IN BIOTECHNOLOGY
2013; 31 (1): 10-19
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
2013; 42 (13): 5788-5808
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
2012; 12 (22): 4884-4893
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
2012; 1 (5): 661-668
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
WILEY-BLACKWELL. 2012: 366–366
View details for Web of Science ID 000308313003155
Release of Magnetic Nanoparticles from Cell-Encapsulating Biodegradable Nanobiomaterials
2012; 6 (8): 6640-6649
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
2012; 7 (4): 553-564
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
2012; 1 (2): 149-158
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
2012; 12 (8): 1508-1515
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
Portable microfluidic chip for detection of Escherichia coli in produce and blood
INTERNATIONAL JOURNAL OF NANOMEDICINE
2012; 7: 2591-2600
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
Simple filter microchip for rapid separation of plasma and viruses from whole blood
INTERNATIONAL JOURNAL OF NANOMEDICINE
2012; 7: 5019-5028
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
The assembly of cell-encapsulating microscale hydrogels using acoustic waves
2011; 32 (31): 7847-7855
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 2011; 23 (37): 4254-4260
Microengineering methods for cell-based microarrays and high-throughput drug-screening applications
2011; 3 (3)
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
2011; 99 (6)
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
2011; 6 (6): 1115-1129
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
2011; 6 (7)
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
2011; 6 (7)
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
2011; 5 (2)
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
2011; 17 (6): 641-649
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
2011; 6 (4)
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
2011; 6 (3)
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
2011; 6 (3)
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
2011; 6 (2): 204-212
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
2011; 6 (2): 138-149
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
2011; 11 (20): 3411-3418
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
2011; 11 (15): 2535-2540
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
2011; 11 (23): 3979-3989
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
2010; 28 (6): 770-781
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
2010; 22 (8)
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
2010; 106 (1): 138-148
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
2010; 5 (3): 469-484
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
Vitrification and levitation of a liquid droplet on liquid nitrogen
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
2010; 107 (10): 4596-4600
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
2010; 2 (1)
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
2010; 368 (1912): 561-583
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
2010; 16 (1): 157-166
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
2009; 25 (1): 253-258
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
2009; 395 (1): 185-193
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
2009; 24 (11): 3208-3214
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
Cell Proliferation in Bioprinted Cell-Laden Collagen Droplets
35th Annual Northeast Bioengineering Conference
IEEE. 2009: 390–391
View details for Web of Science ID 000268886300192
Rapid automated cell quantification on HIV microfluidic devices
LAB ON A CHIP
2009; 9 (23): 3364-3369
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
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
IEEE. 2009: 366–367
View details for Web of Science ID 000268886300180
Ultra wide-field lens-free monitoring of cells on-chip
LAB ON A CHIP
2008; 8 (1): 98-106
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
- Rewritable self-assembled long-period gratings in photonic bandgap fibers using microparticles OPTICS COMMUNICATIONS 2007; 270 (2): 225-228
- Acoustic picoliter droplets for emerging applications in semiconductor industry and biotechnology JOURNAL OF MICROELECTROMECHANICAL SYSTEMS 2006; 15 (4): 957-966
- Droplet-based photoresist deposition APPLIED PHYSICS LETTERS 2006; 88 (14)
- Direct etch method for microfludic channel and nanoheight post-fabrication by picoliter droplets APPLIED PHYSICS LETTERS 2006; 88 (5)
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
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 2005; 18 (4): 709-715
- Picolitre acoustic droplet ejection by ferntosecond laser micromachined multiple-orifice membrane-based 2D ejector arrays ELECTRONICS LETTERS 2005; 41 (22): 1219-1220
- Picoliter droplets for spinless photoresist deposition REVIEW OF SCIENTIFIC INSTRUMENTS 2005; 76 (6)
Coherent array imaging using phased subarmys. Part II: Simulations and experimental results
IEEE TRANSACTIONS ON ULTRASONICS FERROELECTRICS AND FREQUENCY CONTROL
2005; 52 (1): 51-64
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 2004; 17 (4): 517-524
Forward-viewing CMUT arrays for medical Imaging
IEEE TRANSACTIONS ON ULTRASONICS FERROELECTRICS AND FREQUENCY CONTROL
2004; 51 (7): 887-895
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
IEEE. 2003: 1875–1878
View details for Web of Science ID 000189492100435
2D acoustically actuated micromachined droplet ejector array
IEEE International Ultrasonics Symposium
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
2002; 49 (11): 1596-1610
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
ELSEVIER SCIENCE BV. 2002: 471–76
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
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
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
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
IEEE. 2001: 957–960
View details for Web of Science ID 000176890800203