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A guide to light-sheet fluorescence microscopy for multiscale imaging

Abstract

The impact of light-sheet fluorescence microscopy (LSFM) is visible in fields as diverse as developmental and cell biology, anatomical science, biophysics and neuroscience. Although adoption among biologists has been steady, LSFM has not displaced more traditional imaging methods despite its often-superior performance. One reason for this is that the field has largely conformed to a do-it-yourself ethic, although the challenges of big image data cannot be overstated. With the most powerful implementations of LSFM available to only a few groups worldwide, the scope of this technique is unnecessarily limited. Here we elucidate the key developments and define a simple set of underlying principles governing LSFM. In doing so, we aim to clarify the decisions to be made for those who wish to develop and use bespoke light-sheet systems and to assist in identifying the best approaches to apply this powerful technique to myriad biological questions.

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Figure 1: Light-sheet fluorescence microscopy.
Figure 2: Parallelization of light-sheet generation.
Figure 3: Spatial resolution in light-sheet fluorescence microscopy.
Figure 4: Reflected light-sheet geometries.
Figure 5: Gaussian and Bessel beams for light-sheet generation.
Figure 6: Axially swept light-sheet geometries.
Figure 7: Light-sheet penetration.
Figure 8: Multiview imaging.
Figure 9: Ultrafast volumetric imaging.
Figure 10: Hyperspectral light-sheet microscopy.

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Acknowledgements

This work was supported by the Max Planck Society (R.M.P. and J.H.), the European Research Council (ERC Consolidator grant 647885; J.H.) and a fellowship from the Human Frontier Science Program (HFSP) (LT000321/2015-C; R.M.P.). We thank members of the Huisken lab for discussions and critical comments on the manuscript.

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Correspondence to Jan Huisken.

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J.H. is a co-inventor on patent US 20060033987 and an inventor on patent US 20110115895, which are related to light-sheet microscopy.

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Power, R., Huisken, J. A guide to light-sheet fluorescence microscopy for multiscale imaging. Nat Methods 14, 360–373 (2017). https://doi.org/10.1038/nmeth.4224

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