Epr8208

Total proteins were extracted from the ileum tissue and IPI-2I cells following the procedure described previously (22 (link)). Subsequently, western blotting was performed following the standard protocol (22 (link)). The antibodies used were: Occludin (1:1,000; EPR8208, Abcam), Bax (1:5,000; 50599-2-lg, Proteintech), Bcl-2 (1:1,000; 12789–1-AP, Proteintech), β-Actin (1:1,000; 4970, CST), HRP Conjugated AffiniPure Goat Anti-Rabbit IgG (1:5,000; BA1055, BOSTER).

A single-cell suspension was prepared by cutting and disrupting one representative intestinal sample from each group through a 70-μm filter membrane. Cells were diluted with staining buffer (1 × 10⁶ cells/mL) after blockage with 3% FBS. For analysis, the cells were incubated with an anti-occludin antibody (EPR8208, Abcam, United States), anti-ZO-1 antibody (21773-1-AP, Proteintech, United States) and an anti-caspase antibody (25546-1-AP, Proteintech, United States) for 2 h, and then they were incubated with fluorescein isothiocyanate (FITC, SA00003-2) in the dark for 1 h. Occludin, ZO-1, and caspase-3 expression was measured using flow cytometry (FACSCalibur, Becton, Dickinson, and Company, United States).