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Prostate tissues from C57BL/6 mice were cleared and labeled by the anti-GFP antibody for eYFP staining following a modified iDISCO protocol (Renier et al., 2014 (link)) by using a milder index-matching reagent ethyl cinnamate. Cleared and labeled specimens were placed in custom 3D machined sample holders as described previously (Glaser et al., 2019 (link)). The samples were imaged on a Lightspeed Microscopy open-top light-sheet microscope with 20× magnification (0.44 microns per pixel) using a multi-immersion objective (#54–10–12, Special Optics, distributed by Applied Scientific Instrumentation). The fluorescence was filtered with band-pass filters for the 488 nm (for eYFP and EdU staining; FF03–525/50–25, Semrock) and 638 nm (for TOPRO3 nuclear staining; FF01–721/65–25, Semrock) excitation wavelengths. The raw image files were aligned and stitched in ImageJ using BigStitcher software (Hörl et al., 2019 (link)) and fused to TIFF files. The resulting TIFF files were visualized and analyzed using Aivia software (Aivia v8.5, DRVision Technologies LLC, Bellevue, WA). Aivia Pixel Classifier tool was used to segment and enumerate individual nuclei, eYFP and EdU signals.