Goat anti rabbit igg h l hrp pre adsorbed antibody

Total proteins were extracted from BEAS-2B cells following treatment with RIPA lysis buffer (Shanghai Absin Biotechnology Co., Ltd.) on ice. The cells were centrifuged at 16,000 x g for 10 min at 4˚C, and the supernatant was collected for western blotting. A bicinchoninic acid protein assay kit (Thermo Fisher Scientific, Inc.) was used to detect the protein concentrations. Subsequently, protein extracts (30 µg) were separated by 10% SDS-PAGE and were then transferred onto PVDF membranes (Sigma-Aldrich; Merck KGaA). Following the block with 5% non-fat milk for 60 min at room temperature, PVDF membranes were incubated with primary antibodies Bcl-2 (1:1,000; product code ab32124), Bax (1:1,000; product code ab32503), PPARγ (1:1,000; product code ab272718), phosphorylated (p)-NF-κB p65 (1:1,000; product code ab32536), NF-κB p65 (1:1,000; product code ab207297) or GAPDH (1:1,000; product code ab9485) at 4˚C overnight. Subsequently, the membranes were cultivated with goat anti-rabbit IgG H&L (HRP) preadsorbed antibody (1:5,000; product code ab7090; all from Abcam) at room temperature for 1 h. An ECL system (Beyotime Institute of Biotechnology) was used to develop the membranes and blots were analyzed using ImageJ 1.8.0 (National Institutes of Health).