Rabbit monoclonal anti snail antibody

Cell lysates were harvested using RIPA lysis buffer for 30 mins on ice and centrifuged at 13,000 rpm for 10 mins at 4°C. Protein concentration of the supernatant was determined by Bio-Rad Protein Assay (Bio-Rad Laboratories, Inc., USA). An equal amount of each protein extract (30 μg) was resolved using 10% polyacrylamide gel and electro-transferred onto 0.45 μm hybridization nitrocellulose filter (HATF) membrane (Millipore, USA) using Trans-blot Turbo (Bio-Rad Laboratories, Inc., USA). Membranes were immunoblotted with either rabbit polyclonal anti-IFITM1 antibody (GeneTex, USA), rabbit polyclonal anti-actin antibody (Abcam, USA), rabbit monoclonal anti-SNAIL antibody (Cell Signaling, USA), rabbit monoclonal anti-E-Cadherin antibody (Cell Signaling, USA), mouse monoclonal anti-Fibronectin antibody (Abcam, USA), or mouse monoclonal anti-N-Cadherine antibody (BD, USA) overnight at 4°C. Membranes were incubated with either HRP-conjugated anti-rabbit immunoglobulin (Cell Signaling, USA) or HRP-linked anti-mouse immunoglobulin (Cell Signaling, USA) for 1 hr at room temperature. The protein signal was detected by enhanced chemiluminescence (Thermo, USA) using the Amersham Imager 600 (GE Healthcare Life Sciences, UK).