Anti e cadherin antibody 20874 1 ap

Cells were disrupted by RIPA Lysis Buffer (Beyotime) and scraped off the plate surface with a cell scraper. Protein concentration was measured using an enhanced BCA Protein Assay Kit (Beyotime). SDS-PAGE (10%) gels were prepared using SDS-PAGE Gel Quick Preparation Kit (Beyotime). Equal amounts of protein by weight were loaded on the SDS-PAGE and electrophoresed at 80 V for 30 minutes followed by 120 V for 45 minutes. Subsequently, the proteins on the gel were transferred on to polyvinlyidene difluoride membranes (EMD Millipore, Billerica, MA, USA). Membranes were blocked with 5% skim milk for 2 hours, and incubated with primary antibodies overnight at 4°C. Following incubation with the corresponding HRP-conjugated secondary antibody for 1 hour, membranes were developed with SuperSignal West Femto Maximum Sensitivity Substrate (Thermo Fisher Scientific). GAPDH was used as an internal loading control. The following antibodies were used: anti-LIPH antibody (Abcam, ab192615), anti-HIF-1α antibody (36169s; CST; Boston, MA, USA), anti-Slug antibody (12129-1-AP; Proteintech Group), anti-E-cadherin antibody (20874-1-AP; Proteintech Group), anti-N-cadherin antibody (22018-1-AP; Proteintech Group), and HRP-conjugated AffiniPure goat anti-rabbit IgG(H+L) (SA00001-2; Proteintech Group).