TUNEL Staining for Apoptosis Detection

Apoptosis‐positive cells in the rat brain group were detected using TUNEL staining kits according to the manufacturer's instructions (Servicebio, G1507). Brain sections of each group were routinely dewaxed for 10 min and dehydrated with gradient alcohol. Then the sections were addressed with proteinase K (20 mg/L) for 15 min and 0.5% H₂O₂ for 20 min. After PBS washing, the sections were incubated with TUNEL (50 μL) for 60 min in a wet chamber at 37°C. Then the sections were disposed of fluorescein antibody. Then the sections were subjected to multiple procedures including diaminobenzidine (DAB) incubation, hematoxylin treatment, xylene, gradient ethanol dehydration, and neutral resin sealing. An inverted microscopy was adopted to obtain tissue images (Feng et al., 2020 (link)).