P53 antibody 9282

Twenty μg of protein per sample were loaded in SDS-PAGE in 10% (for E2F1,Chk1 and p53) or 8% (for Rb or PARP-1) polyacrylamide gels, and then transferred onto nitrocellulose membranes. Antibody dilutions were as follows: Chk1- 1:500 (sc-377231), E2F1- 1:1000 (sc-193), Rb- 1:200 (sc-102), PARP-1- 1:1000 (H-300: sc-25780), Cleaved Caspase-3 (Asp175) Antibody #9661 1:1000, p53 Antibody #9282 1:1000 (Cell signaling) in 5% fat free milk 0’05% TTBS. HA Antibody- 1:2000 (Boehringer mannheim), Flag antibody- 1:2000 (Sigma)

Between April 2007 and April 2008, we recruited 146 patients with primary GC from the Zhongshan hospital and Shanghai Cancer Center, Fudan University. All the patients (excluding the stage IV disease) underwent an R0 resection (with no tumour cells at the margin) with D2 lymph node dissection, and all the patients histopathologically confirmed for the diagnosis of stages II and III received 6–8 courses of chemotherapy-based fluorouracil following the operation. The Ethics Committee of Fudan University approved our study. All the participants provided written informed consent prior to enrolment. The human GC cell lines were obtained from the American Type Culture Collection (Manassas, VA, USA) and were identified through an STR analysis. The p53 antibody (#9282) and p21 antibody (#8831) were purchased from Cell Signaling Technology (Beverly, MA, USA). Nutlin-3 (S1061) was purchased from Selleck (Princeton, NJ, USA).