Anti phospho met antibody

Cell lysates were extracted with a lysis buffer (20 mM Tris-HCl (pH = 8.0; FUJIFILM Wako), 150 mM NaCl (FUJIFILM Wako), 2 mM EDTA (FUJIFILM Wako), 100 mM NaF (FUJIFILM Wako), 1% NP40 (FUJIFILM Wako), 1 μg/mL leupeptin (Sigma), 1 μg/mL antipain (Sigma), and 1 mM phenylmethylsulfonyl fluoride (Sigma)). Phosphorylated protein and total protein in these lysates were examined by western blotting assay by using following primary antibodies: anti-phospho-Met (Tyr1234/1235) antibody, anti-phospho-Met antibody (Tyr1349), anti-Met antibody, anti-phospho-ERK1/2 antibody, anti-ERK1/2 antibody, anti-phospho-Akt antibody, anti-Akt antibody, anti-phospho-p38MAPK antibody, anti-p38MAPK antibody, anti-phospho-NF-κB antibody, anti-NF-κB antibody (Cell Signaling Technology, Beverly, MA, USA), anti-RANKL antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and anti-β-actin antibody (Sigma). Proteins were visualized using Luminata Forte (Merck Millipore, Nottingham, UK) according to the manufacturer’s instructions.

Anti-SAAL1 antibody and anti-Lamin B2 antibody were purchased from Bethyl and Abcam. Anti-phospho-Met antibody, anti-Met antibody, anti-phosphor-mTOR antibody, anti-mTOR antibody, anti-phosphor-Akt antibody, anti-Akt antibody, and anti-Na and K-ATPase antibody were purchased from Cell Signaling Technology. Anti-α-Tubulin antibody, anti-Actin antibody, and the HRP-conjugated secondary antibodies were purchased from Santa Cruz Biotechnology Inc. Sorafenib and foretinib were purchased from Merk and Selleckchem, respectively. The detailed information on antibodies and reagents are listed in the Table S2.