Resonant Electro-Optic Imaging for Microscopy at Nanosecond Resolution.
We demonstrate an electro-optic wide-field method to enable fluorescence lifetime microscopy (FLIM) with high throughput and single-molecule sensitivity. Resonantly driven Pockels cells are used to efficiently gate images at 39 MHz, allowing fluorescence lifetime to be captured on standard camera sensors. Lifetime imaging of single molecules is enabled in wide field with exposure times of less than 100 ms. This capability allows combination of wide-field FLIM with single-molecule super-resolution localization microscopy. Fast single-molecule dynamics such as FRET and molecular binding events are captured from wide-field images without prior spatial knowledge. A lifetime sensitivity of 1.9 times the photon shot-noise limit is achieved, and high throughput is shown by acquiring wide-field FLIM images with millisecond exposure and >108 photons per frame. Resonant electro-optic FLIM allows lifetime contrast in any wide-field microscopy method.
View details for DOI 10.1021/acsnano.1c04470
View details for PubMedID 34546704
- Fast pulse shaping for a novel gated electron mirror REVIEW OF SCIENTIFIC INSTRUMENTS 2021; 92 (4)
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Design for a 10keV multi-pass transmission electron microscope.
2019; 207: 112834
Multi-pass transmission electron microscopy (MPTEM) has been proposed as a way to reduce damage to radiation-sensitive materials. For the field of cryo-electron microscopy (cryo-EM), this would significantly reduce the number of projections needed to create a 3D model and would allow the imaging of lower-contrast, more heterogeneous samples. We have designed a 10keV proof-of-concept MPTEM. The column features fast-switching gated electron mirrors which cause each electron to interrogate the sample multiple times. A linear approximation for the multi-pass contrast transfer function (CTF) is developed to explain how the resolution depends on the number of passes through the sample.
View details for DOI 10.1016/j.ultramic.2019.112834
View details for PubMedID 31520925
Electro-optic imaging enables efficient wide-field fluorescence lifetime microscopy.
2019; 10 (1): 4561
Nanosecond temporal resolution enables new methods for wide-field imaging like time-of-flight, gated detection, and fluorescence lifetime. The optical efficiency of existing approaches, however, presents challenges for low-light applications common to fluorescence microscopy and single-molecule imaging. We demonstrate the use of Pockels cells for wide-field image gating with nanosecond temporal resolution and high photon collection efficiency. Two temporal frames are obtained by combining a Pockels cell with a pair of polarizing beam-splitters. We show multi-label fluorescence lifetime imaging microscopy (FLIM), single-molecule lifetime spectroscopy, and fast single-frame FLIM at the camera frame rate with 103-105 times higher throughput than single photon counting. Finally, we demonstrate a space-to-time image multiplexer using a re-imaging optical cavity with a tilted mirror to extend the Pockels cell technique to multiple temporal frames. These methods enable nanosecond imaging with standard optical systems and sensors, opening a new temporal dimension for wide-field low-light microscopy.
View details for DOI 10.1038/s41467-019-12535-5
View details for PubMedID 31594938