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Reflective imaging improves spatiotemporal resolution and collection efficiency in light sheet microscopy.


ABSTRACT: Light-sheet fluorescence microscopy (LSFM) enables high-speed, high-resolution, and gentle imaging of live specimens over extended periods. Here we describe a technique that improves the spatiotemporal resolution and collection efficiency of LSFM without modifying the underlying microscope. By imaging samples on reflective coverslips, we enable simultaneous collection of four complementary views in 250?ms, doubling speed and improving information content relative to symmetric dual-view LSFM. We also report a modified deconvolution algorithm that removes associated epifluorescence contamination and fuses all views for resolution recovery. Furthermore, we enhance spatial resolution (to <300?nm in all three dimensions) by applying our method to single-view LSFM, permitting simultaneous acquisition of two high-resolution views otherwise difficult to obtain due to steric constraints at high numerical aperture. We demonstrate the broad applicability of our method in a variety of samples, studying mitochondrial, membrane, Golgi, and microtubule dynamics in cells and calcium activity in nematode embryos.

SUBMITTER: Wu Y 

PROVIDER: S-EPMC5682293 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Reflective imaging improves spatiotemporal resolution and collection efficiency in light sheet microscopy.

Wu Yicong Y   Kumar Abhishek A   Smith Corey C   Ardiel Evan E   Chandris Panagiotis P   Christensen Ryan R   Rey-Suarez Ivan I   Guo Min M   Vishwasrao Harshad D HD   Chen Jiji J   Tang Jianyong J   Upadhyaya Arpita A   La Riviere Patrick J PJ   Shroff Hari H  

Nature communications 20171113 1


Light-sheet fluorescence microscopy (LSFM) enables high-speed, high-resolution, and gentle imaging of live specimens over extended periods. Here we describe a technique that improves the spatiotemporal resolution and collection efficiency of LSFM without modifying the underlying microscope. By imaging samples on reflective coverslips, we enable simultaneous collection of four complementary views in 250 ms, doubling speed and improving information content relative to symmetric dual-view LSFM. We  ...[more]

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