High resolution whole brain neuron network imaging by orthogonal light sheet microscopy with focus sweeping

摘要

FxClear, a free-hydrogel electrophoretic tissue clearing method for rapid de-lipidation of tissues with high preservation of immunoreactivity Over the last two decades, several tissue clearing methodolo- gies have been established that render tissues optically transparent and allow imaging of unsectioned tissues of significant volumes, thus improving the capacity to study the relationships between cell and 3D tissue architecture. Despite these technical advances, the important unsolved challenges that these methods face include complexity, time, consistency of tissue size before and after clear- ing, and ability to immunolabel various antibodies in cleared tissue. Here, we established very simple and fast tissue clearing proto- col, FxClear, which involves acrylamide-free electrophoretic tissue clearing (ETC). By removal of the acrylamide infusion step, we were able to achieve fast reaction time, smaller tissue expan- sion, and higher immunoreactivity. Especially, immunoreactivity and fluorescence intensity were increased in FxClear-processed tissues compared to un-cleared tissues. Our protocol may be suitable for small-sized biopsy samples for 3D pathological examinations.