Rabbit polyclonal antibody against p38

Proteins were extracted by lysing cells in sodium dodecyl sulfate (SDS) sample buffer (62.5 mM Tris–HCl [pH 6.8], 2% SDS) containing 1 mM phenylmethylsulfonyl fluoride, 10 μg/mL pepstatin, 12.5 μg/mL leupeptin, 2 μg/mL aprotinin, 1 mM sodium orthovanadate, and 1 mM sodium molybdate. Cell extracts were processed for western immunoblotting as described previously [50 (link)]. The following antibodies used for immunoblotting were purchased from the indicated suppliers: mouse monoclonal antibody against AF1q from Abnova (Taipei City, Taiwan), mouse monoclonal antibody against GAPDH from Sigma Aldrich (St Louis, MO), mouse monoclonal antibody against cytokeratin 8/18 from Santa Cruz Biotechnology (Santa Cruz, CA), rabbit polyclonal antibody against Fibronectin from Sigma Aldrich, mouse monoclonal antibody against Vimentin from Abcam (Cambridge, UK), rabbit polyclonal antibody against p38 from Santa Cruz Biotechnology, rabbit polyclonal antibody against Thr180/Tyr182 p38 phosphorylations from Cell Signaling (Danvers, MA), rabbit polyclonal antibody against Erk1/2 from Sigma Aldrich, rabbit polyclonal antibody against Thr202/Tyr204 Erk1/2 phosphorylations from Cell Signaling.

Gemcitabine (GEM) was purchased from Eli Lilly (Indianapolis, IN, USA). AMD3100, MTT, Hoechst 33342, PD98059 (an ERK inhibitor), PF573228 (a FAK inhibitor), SB20938 (a P38 inhibitor) were all supplied by Sigma-Aldrich (St. Louis, MO, USA). Recombinant human SDF-1α (rhSDF-1α), the anti-IL-6 mouse monoclonal antibody (mAb) and the IL-6 ELISA kit were purchased from R&D Systems (Minneapolis, MN, USA). The anti-αSMA mouse mAb was from Dako. The anti-vimentin and anti-SDF-1α mouse mAbs, the anti-CXCR4 rabbit polyclonal antibody and the SDF-1α neutralizing antibody (SDF-1α Ab) were from Abcam. Rabbit mAbs against pAKT (Ser473), pP38 (Thr180/Tyr182), ERK1/2, pERK1/2 (Thr202/Tyr204) and pFAK (Tyr397) were from Cell Signaling Technologies (Beverly, MA, USA). The mouse mAb against FAK, rabbit mAbs against AKT and and β-actin, the rabbit polyclonal antibody against P38, and the secondary horseradish peroxidase-conjugated anti-rabbit and anti-mouse antibodies were all purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).