Enhanced chemiluminescence ecl western blotting detection reagent

Human colorectal carcinoma cell line, RKO, was obtained from American Type Culture Collection (Manassas, Virginia, USA). Minimum essential medium (MEM) was purchased from Gibco Laboratories (Paisley, UK). Fetal bovine serum (FBS) and [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] (MTT) were purchased from Sigma-Aldrich Chemical Company (Dorset, UK). Six-well flat-bottom tissue culture plates were purchased from Nunc Inc. (Hereford, UK). Phenylmethylsulfonyl fluoride (PMSF) was purchased from Sigma-Aldrich Co., Ltd. (St. Louis, MO, USA) and protease inhibitor mixture solution was obtained from Wako Pure Chemical Industries, Ltd. (Osaka, Japan). Enhanced chemiluminescence (ECL) western blotting detection reagents and pure nitrocellulose MEMbranes were purchased from GE Healthcare (Buckinghamshire, UK). Primary antibodies for the detection of caspase 3 and β-actin and goat antirabbit and goat anti-mouse IgG horseradish peroxidase (HRP)-conjugated secondary antibodies were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). All chemicals and reagents were of analytical grade and purchased from Sigma-Aldrich Co., Ltd. unless otherwise specified.