Hepatocellular carcinoma (HCC), among the most common malignancies worldwide, remains a major threat to public health, and there is an urgent need to identify novel biomarkers for diagnosis, prognosis and targets for anti-cancer treatment. In this study, two-dimensional polyacrylamide gel electrophoresis coupled with ESI-Q-TOF MS/MS analysis was used to identify differentially expressed proteins among the HCC tumour centre, tumour margin and nontumourous liver tissues. In total, 52 spots with significant alteration were positively identified by MS/MSanalysis. Altered expression of representative proteins, including CIB1, was validated by Western blotting. Immunostaining suggested an increase tendency of CIB1 expression from nontumourous liver tissue to tumour centre. Knockdown of CIB1 expression by RNA interference led to the significant suppression of the cell growth in hepatoma HepG2 cells. These data suggest that CIB1 may be used as a novel prognostic factor and possibly an attractive therapeutic target for HCC.