Ab17185

Proteins were extracted from cells or tissues and added to the RIPA buffer. Cracking on ice for 30_60 minutes. The supernatant was collected by centrifugation at 4°C and 12000 rpm for 15 minutes. BCA protein assay kit (Boster Inc, China) was used for protein quantification. SDS-PAGE gel electrophoresis was performed. Then, the protein was transferred to the nitrocellulose membrane and 5% skimmed milk powder blocked the nonspecific antigen. The primary antibody was added and incubated at 4°C overnight. After washing the membrane, horseradish peroxidase-labeled IgG (secondary antibody) was added at 37 °C for 1h. Densitometric analysis was performed by Image Lab. In the assay, antibodies included rabbit anti-hnRNP E1 (Abcam, UK. ab74793, 1:1000), mouse anti-HPV16 E2 (Abcam, UK. ab17185, 1:1000), mouse anti-HPV16 E6 (Abcam, UK. Ab70, 1:1000), mouse anti-β-actin (Boster Inc, China, 1:300).

Heart samples were drawn at 2 h after reperfusion and detected as previously described [17 (link)]. Frozen heart tissue was homogenized in lysis buffer. The protein concentration was determined using a BCA Protein assay kit (Cwbiotech, China). Next, 2 µg/µl of protein was separated using 10% SDS-PAGE and then electrophoretically transferred onto PVDF membranes (Millipore, USA). Proteins on the membranes were then probed using primary antibodies, including mouse Bcl₂ (ab7973, abcam), Bax (ab32503, abcam), Caspase-3 (ab17185, abcam), NF-κBp65 (ab16502, abcam), and GAPDH (5174, CST). Following incubation with secondary antibodies, including goat anti-rabbit secondary antibody, the positive protein blots were developed using a chemiluminescent system, and the bands were quantified using the Gel Image system ver. 4.00 (Tanon, China). Data were shown as the percentage density of the blots.