Multi analyst

Cells were lysed on ice for 30 min using RIPA buffer followed by high-speed centrifugation. Afterwards, total cell protein concentration was determined using a BCA Protein Assay Kit (Thermo Fisher Scientific, USA). After blocking, membranes were incubated with antibodies against the desired primary antibodies including Ki67 (ab16667, abcam, USA), E-cadherin (ab40772, abcam, USA), N-cadherin (ab245117, abcam, USA), and GAPDH (10494-1-AP, proteintech, USA). Bands were visualized using enhanced chemiluminescence reagents (NCM, USA) and analyzed using a gel documentation system (Bio-Rad Gel Doc1000 and Multi-Analyst version 1.1). Relative band intensity was determined by densitometry software and normalized with GAPDH protein.

Cells were lysed on ice for 30 min with lysis buffer. Total cell protein concentration was determined using a BCA Protein Assay Kit. The total protein (30 μg) was resolved using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Invitrogen) and transferred to a polyvinylidene fluoride membrane (Roche). Membranes were blocked with 5% bovine serum albumin in TBS-Tween (0.1%) and incubated against the desired antibodies. The primary antibodies BAX (5023, Cell Signaling Technology, USA), BCL-2 (ab196495, abcam, USA), p53 (21083, Signalway Antibody, USA), AMP-activated protein kinase (AMPK, 5831, Cell Signaling Technology), phosphorylation of AMPK (p-AMPK, 500815, Cell Signaling Technology), XIAP (14334, Cell Signaling Technology), GAPDH (5174, Cell Signaling Technology), TSG101 (14497, Proteintech, United States), CD63 (25682, Proteintech), CD81 (66866, Proteintech), and horseradish peroxidase-conjugated secondary antibody (Santa Cruz) were used. Bands were visualized using enhanced chemiluminescence reagents and analyzed using a gel documentation system (Bio-Rad Gel Doc1000 and Multi-Analyst version 1.1).