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We developed a Bessel light sheet fluorescence microscopy (LSFM) system toenable high-speed, wide-field intra-vital imaging of zebrafish and other thickbiological samples. This system uses air objectives for the convenient mountingof large samples and incorporates an electrically tunable lens for automaticfocusing during volumetric imaging. To enhance the precision of 3D imaging, theimpact of the electrically tunable lens on system magnification is investigatedand modified through designed experiments. Despite using Bessel beams with sidelobes, we achieved satisfactory image quality through a straightforwardbackground noise subtraction method, eliminating the need for furtherdeconvolution. Our system provides zebrafish imaging at a resolution comparableto commercial confocal microscopy but in just 1/40th of the time. We alsointroduced light field microscopy (LFM) to improve 3D in vivo imaging temporalresolution. Apart from the 28-fold speed enhancement, the comparison of LFM andLSFM results reveals a unique aspect of LFM imaging concerning image dynamicrange, which has not been previously reported.

Light sheet and light field microscopy based on scanning Bessel beam illumination