Rabbit anti rbp4 antibody

Ovarian endometriosis tissues and normal eutopic endometrium tissues were lysed in RIPA buffer containing protease and phosphatase inhibitor cocktail. After centrifugation at 12,000 rpm and 4°C for 15 min, the protein concentration in the supernatant was determined using a BCA protein assay. Equal amounts of total protein (30 µg) were loaded and separated on 12.5% SDS-PAGE and then transferred to PVDF membranes. After blocking with protein-free rapid blocking buffer for 0.5 h at room temperature, the membranes were incubated with primary antibodies overnight at 4°C. Rabbit anti-RBP4 antibody (1:5000; Abcam) and rabbit anti-β-actin (1:1000, cell signaling technology) were used as primary antibodies, with β-actin serving as the internal loading control. Then, the membranes were then incubated with secondary antibodies (1:3000) for 1 h at room temperature. Excess secondary antibody was removed by three washes in TBST. The targeted protein bands were visualized and imaged using an ECL Western blotting kit (NCM Biotech, China).