Lattice light-sheet microscopy (LLSM) is a very efficient technique for high resolution 3D imaging of dynamic phenomena in living biological samples. However, LLSM imaging remains limited in depth due to optical aberrations caused by sample-based refractive index mismatch. Here, we propose a simple and low-cost active image optimization (AIO) method to recover high resolution imaging inside thick biological samples. AIO is based on (1) a light-sheet autofocus step (AF) followed by (2) an adaptive optics image-based optimization. We determine the optimum AIO parameters to provide a fast, precise and robust aberration correction on biological samples. Finally, we demonstrate the performances of our approach on sub-micrometric structures in brain slices and plant roots.
We report on an Adaptive Optics (AO) Light-Sheet Fluorescence Microscope compatible with neuroimaging, based on direct wavefront sensing without the requirement of a guide star. We demonstrate fast AO correction, typically within 500ms, of in-depth aberrations of the live adult Drosophila Melanogaster brain, enabling to double the contrast when imaging with structural or calcium sensors. We quantify the gain in terms of image quality on multiply neuronal structures part of the sleep network in the Drosophila brain, at various depths, and discuss the optimization of key parameters driving AO such as the number of corrected modes and the photon budget. We present a first design of a compact AO add-on that is compatible with integration into most of reported Light-Sheet setups and neuroimaging.
We present a novel implementation of extended-scene adaptive optics for light-sheet microscopy based on direct wavefront sensing for fast aberration correction. We report AO-enhanced functional images of GCaMP neurons in the live drosophila brain.
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