Andgoat anti rabbit

Six days after lentiviral infection, SMMC-7721 cells were collected, washed with ice-cold PBS and lysed in 2× SDS sample buffer (100 mM Tris-Hcl (pH 6.8), 10 mM EDTA, 4% SDS, 10% Glycine). Equal amount of protein samples (30 μg) were run on the 10% SDS-PAGE gel at 50 V for 3 h. The proteins were then transferred to polyvinylidene fluoride (PVDF) membrane at 300 mA for 1.5 h. The membrane was blocked by 1% bovine serum albumin (BSA) in TBST at RT for 1 h and incubated with primary antibodies, mouse anti-USP39 (1:100, Sigma Aldrich, SAB1407042) , rabbit anti-PARP (1: 1000, Cell Signaling Technology, #9542), rabbit anti-Caspase 3 (1: 500, Cell Signaling Technology, #9661), rabbit anti- Caspase 9 (1:1000, Proteintech Group Inc., 10380-1-AP), rabbit anti-Bax (1: 500, Cell Signaling Technology, #2774) and rabbit anti-GAPDH (1:100,000, Proteintech Group Inc., 10494-1-AP) overnight at 4°C. After washed by TBST, membranes were incubated with horseradish peroxidase conjugated goat anti-mouse (1:5,000, Santa Cruz, SC-2005) and goat anti-rabbit (1:5,000, Santa Cruz, SC-2054) secondary antibodies for 2 h at room temperature. The membrane was washed by TBST and signals were detected by enhanced chemiluminescence kit (Pierce).