Fluar 10 0.5 objective

Digital camera (HDC-HS900GK) was used to acquire the bright-field images of samples. Confocal fluorescence microscopy (LSM710, Zeiss, Germany), equipped with the Fluar $10\times /0.5$ objective (dry, working distance 2.0 mm) and Plan-Apochromat $20\times /0.8$ objective (dry, working distance 0.55 mm), was used to acquire the GFP fluorescence images of brain sections. Before and after clearing, the fluorescence images were obtained under the same imaging parameters.

To obtain the spatial distribution of the MEPs in skeletal muscles, the cleared muscles were imaged using a light sheet fluorescence microscope (LaVision BioTec I, Bielefeld, Germany) equipped with an sCMOS camera (Andor Neo), a ×2/0.5 objective lens equipped with a dipping cap, and an Olympus MVX10 zoom microscope body (magnification range of ×0.63–×6.3). The cleared tissues were mounted on the sample holder and incubated with DBE in the sample reservoir. The z-step interval was 5 μm.
To analyze the labeled motor neurons or the intramuscular diffusion of the CTB, the whole spinal cord or muscle slices were imaged with inverted confocal fluorescence microscopy (LSM710; Zeiss, Oberkochen, Germany) equipped with Fluar ×5/0.25 objective (dry, W.D. 12.5 mm) and Fluar ×10/0.5 objective (dry, W.D. 2.0 mm). The cleared spinal cord segments or muscle slices were placed on a coverslip with the clearing reagent (DBE) or 0.01 M PBS, with another coverslip being put on the top of the sample.