Horseradish peroxidase conjugated goat anti rabbit igg antibody

Western blotting was performed as described in our previous study (Chen et al., 2019 (link)). Briefly, cells were lysed and samples were prepared using RIPA protein extraction reagent (Beyotime, Shanghai, China) and a protease inhibitor cocktail (Roche, IN, United States) according to the manufacturer’s protocol. Next, samples with equal amounts of protein were electrophoresed on 10% SDS-PAGE gels and then transferred onto PVDF membranes (Millipore, MA, United States). Membranes were blocked in 5% fat-free milk in TBST buffer, then incubated with primary antibodies (GAPDH antibody, 1:5000 dilution, Proteintech; anti-rabbit PYGB antibody, Ki-67 antibody, N-cadherin, E-cadherin, β-catenin, Snail, Slug, and Twist, 1:1000 dilution; Signalway Antibody, TX, United States) at 4°C overnight. Finally, the second antibody (Horseradish peroxidase-conjugated goat anti-rabbit IgG antibody 1:5000 dilution, Beyotime) was used and the signals were visualized using an Odyssey detection system (Li-COR biosciences, NE, United States).