Anti perk1 2 antibody

Total protein was extracted using RIPA lysis buffer (Sigma) containing protease and phosphatase inhibitors (Beyotime, China) and quantified using the BCA kit (Beyotime, China). Proteins (30 mg of total protein) were separated by 12% or 10% SDS-PAGE and transferred to a polyvinylidene difluoride (PVDF) membrane (Millipore, USA). The membranes were blocked in TBST containing 5% nonfat milk, and incubated overnight at 4°C with various primary antibodies, including anti-FGF2 antibody (Absin); anti-GRB2 antibody (Abcam); anti-ERK1/2 antibody (CST); anti-pERK1/2 antibody (CST); anti-Ras antibody(Abcam); anti-β-Actin antibody(CST); anti-GAPDH antibody(Abcam). Next, it was incubated with an HRP-conjugated goat anti-rabbit IgG (CST). The values of band intensities were detected by ECL kit (Beyotime, China).

Western blot analysis was performed using standard procedures. The following primary antibodies were used in the experiments: anti-FOXM1 antibody (Santa Cruz), anti-pAKT antibody (Peprotech, USA), anti-AKT antibody (Cell Signaling Technology, Beverly, MA, USA), anti-pGSK3β antibody (Proteintech) at 1:1000, anti-Snai1 antibody (Cell Signaling Technology), anti-pIGF1R antibody (Abcam, Cambridge, UK), anti-IGF1R antibody (Abcam, Cambridge, UK), anti-E-cadherin antibody (BD Biosciences, USA), anti-Vimentin antibody (Cell Signaling Technology), anti-N-cadherin antibody (Cell Signaling Technology), anti-pERK1/2 antibody (Abcam), anti-ERK1/2 antibody (Abcam), anti-HIF1α antibody (Novus Biologicals, USA), anti-ETS1 antibody (Abcam) and anti-β-actin antibody (Sigma, St. Louis, MO,USA).