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Fast imaging of live organisms with sculpted light sheets.


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Authors

Chmielewski, Aleksander K 
Kyrsting, Anders 
Mahou, Pierre 
Wayland, Matthew T 

Abstract

Light-sheet microscopy is an increasingly popular technique in the life sciences due to its fast 3D imaging capability of fluorescent samples with low photo toxicity compared to confocal methods. In this work we present a new, fast, flexible and simple to implement method to optimize the illumination light-sheet to the requirement at hand. A telescope composed of two electrically tuneable lenses enables us to define thickness and position of the light-sheet independently but accurately within milliseconds, and therefore optimize image quality of the features of interest interactively. We demonstrated the practical benefit of this technique by 1) assembling large field of views from tiled single exposure each with individually optimized illumination settings; 2) sculpting the light-sheet to trace complex sample shapes within single exposures. This technique proved compatible with confocal line scanning detection, further improving image contrast and resolution. Finally, we determined the effect of light-sheet optimization in the context of scattering tissue, devising procedures for balancing image quality, field of view and acquisition speed.

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Keywords

Animals, Cell Nucleus, Embryo, Nonmammalian, Image Processing, Computer-Assisted, Imaging, Three-Dimensional, Microscopy, Fluorescence, Zebrafish

Journal Title

Sci Rep

Conference Name

Journal ISSN

2045-2322
2045-2322

Volume Title

5

Publisher

Springer Science and Business Media LLC
Sponsorship
Medical Research Council (MR/K015850/1)
This work was funded by grants from the Wellcome Trust, the Medical Research Council, the CamBridgeSense network, Carlsberg Foundation, the Alzheimer Research UK Trust and the Biotechnology and Biological Sciences Research Council and the Wolfson Foundation.