Polyvinylidene fluoride membranes pvdf

The primary antibodies against p-ERK1/2, p-JNK, p-p38, ACE, chymase, Ang II and β-actin, as well as peroxidase (HRP)-conjugated secondary antibodies against goat anti-rabbit IgG and goat anti-mouse IgG, were obtained from Genetex (Irvine, CA, USA) or Cell Signaling Technology (Beverly, MA, USA). Chemiluminescence substrates were visualized using enhanced chemiluminescence detection (Western Lightning Plus-ECL; PerkinElmer, Boston, MA, USA) and a luminescence imaging system (LAS-3000; Fuji Film, Stamford, CT, USA). Polyvinylidene fluoride membranes (PVDF) were purchased from Merck (Kenilworth, NJ, USA), and captopril (ACEI; #C4042) and chymostatin (CI; #C7268) were bought from Sigma-Aldrich (St. Louis, MO, USA).

Rabbit EphA2 polyclonal antibodies and mouse β-actin monoclonal antibodies were obtained from Santa Cruz Biotechnology, Inc., (Dallas, TX, USA). Rabbit Akt, phosphor-Akt (p-Akt), p21, cyclin-dependent kinase 2 (CDK2) and Cyclin E monoclonal antibodies, and mouse p27, retinoblastoma protein (Rb), phosphor-Rb, glycogen synthase kinase-3β (GSK-3β) and phosphor-GSK-3β monoclonal antibodies, and the PI3K/Akt signalling pathway small molecule inhibitor, LY294002, were obtained from Cell Signaling Technology, Inc., (Danvers, MA, USA). Paclitaxel was obtained from Bristol-Myers Squibb (New York, NY, USA). The EphA2 cDNA-pEGFP-N1 expression plasmid and pEGFP-N1 vector plasmid were obtained from GeneChem Co., Ltd. (Shanghai, China). Lipofectamine^® 2000 Transfection Reagent and Opti-MEM^® I Reduced-Serum Medium were obtained from Invitrogen Life Technologies (Carlsbad, CA, USA). 10% foetal bovine serum (FBS) and RPMI-1640 medium were obtained from Hyclone Laboratories, Inc. (Logan, UT, USA). Cell Counting kit-8 (CCK-8), 100 IU/ml penicillin, 100 IU/ml streptomycin, Annexin V-fluorescein isothiocyanate, propidium iodide and BeyoECL Plus Detection system were obtained from Beyotime Institute of Biotechnology (Jiangsu, China). Polyvinylidene fluoride membranes (PVDF) were obtained from EMD Millipore (Billerica, MA, USA).