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To observe three dimensional (3D) in vivo imaging of thick samples in real time, a compact and robust optical sectioning microscope system is required. Recently, we employed static light sheet microscopy (LSM) in which light sheet was generated by a cylindrical lens and objective lens to obtain optically sectioned 3D images. Due to the illumination arm and detection arm are orthogonal to each other in LSM, the photo-bleaching effect of imaging can be reduced and increases the scanning rate so that long-term observation can be achieved. However, the multiple optics in the illumination arm of LSM is too bulky. To overcome this problem, in this work, we show a volume holographic technique can be used to reduce the bulky size of the illumination arm of LSM. Here, we report on design, development, and implementation of holographic light sheet microscope (HLSM) system for in vivo imaging of the biological sample. A VHG corresponding to the wavefront of a cylindrical lens and an objective lens is recorded by superposing it with a reference plane wave into phenanthrenquinone poly (methyl-methacrylate) (PQ-PMMA) photopolymer. Implementation of VHG in LSM makes its smaller and more portable. In order to show the performance of our HLSM, we utilized live Caenorhabditis elegans (C. elegans). Its transparent body allows direct observation of subcellular dynamics with fluorescence-tags. We observed Germline-specific green fluorescent protein (GFP)-labeled embryos and oocytes inside the live C. elegans. As a comparison to bright field microscopy, our HLSM system has inherent optical sectioning ability which helps us in acquiring fine image of the live biologic samples with high contrast. Our method has multiple advantages, including low cost, compact size, easy to implement, and wavelength tunable which makes it suitable for direct implementation in the regular commercial microscope for designing new light sheet microscopic geometries.
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