Anti her3 antibody

Erlotinib was kindly provided by F. Hoffman-La Roche Ltd; TPX-0005 was from TP Therapeutics, Inc.; dasatinib was purchased from Bio Vision; sorafenib was from Toronto Research Chemicals Inc; the other inhibitors were from Selleck Chemicals. Anti-HER2 and anti-pHER2 antibodies were purchased from Upstate Biotechnology; anti-EGFR, anti-pEGFR, anti-pHER3, anti-MET anti-pMET, anti-ERK1/2, anti-pERK1/2, anti-AKT, anti-pAKT (T308 or S473), anti-STAT3, anti-pSTAT3, anti-PTEN, anti-AXL, anti-CDCP1, anti-pCDCP1, anti-SRC, anti-FYN, anti-LYN, anti-YES, anti-LCK, anti-pSFK (Y416), anti-β-catenin, anti-S6K, and anti-pS6K antibodies were from Cell Signaling Technology; anti-E-cadherin antibody was from BD biosciences; anti-HER3 antibody was from Santa Cruz Biotechnology; anti-β-actin antibody was from Abcam, Inc.; Anti-GAPDH antibody was from PROMEGA; α-tubulin antibody was sourced from Sigma-Aldrich.

BT474M1 or rBT474 tumors were collected when mice were sacrificed, fixed with 10% neutral-buffered formalin, and embedded to paraffin. Tissue sections of 4 μm thick were deparaffinized, and heat-induced antigen retrieval was performed in sodium citrate buffer for 20 min. After blocking endogenous peroxidase activity with 3% H₂O₂, 10% normal horse serum was applied for the blocking of nonspecific binding sites. Anti-HER3 antibody (Santa Cruz) was put on the sections and incubated overnight at 4 °C. After washing with PBS, biotinylated secondary antibody (Bio-Rad) was applied for 30 min, followed by an VECTASTAIN ABC kit (Vector Lab) and then the color was developed using the DAB Peroxidase substrate kit (Vector Lab). Counterstaining was performed with hematoxylin.