TUNEL assay for apoptosis in intestine

The terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick-end labelling (TUNEL) assay (4810-30-K, Roche, Beijing Zhongshan Gold Bridge Co., Ltd., Beijing, China) was performed according to the manufacturer’s instructions. Sections were incubated in TUNEL reaction solution for 60 min at 37°C, after being washed 3 times by PBS, and were then incubated in POD for 30 min at 37°C, after being washed 3 times by PBS. Then, 50-100 μL DAB was used for staining and imaging in 15-25°C for 10 min; haematoxylin redyeing. The positive cell data were counted from top to half of the intestinal villus. In total, one hundred thirty-five intestinal villi were counted for each group.