Bax-interacting factor-1 (Bif-1) interacts with Beclin1 [the mammalian ortholog of yeast autophagy-related gene 6 (Atg6)] and affects the formation of autophagosomes during autophagy. The aim of this study was to explore Bif-1 expression and its prognostic significance in comparison with various clinicopathological predictors of survival. Bif-1 protein expression was determined by immunohistochemistry in 206 hepatocellular carcinomas. Cytoplasmic immunoreactivity was scored semi-quantitatively. The results were analyzed in correlation with various clinicopathological characteristics, including patient survival. The Chi-square test and Kaplan-Meier survival analysis were applied. The expression of Bif-1 was significantly higher in the hepatocellular cancers than in the adjacent matched non-tumor tissues (51.5 vs. 33.0%, P<0.01). Increased expression of Bif-1 in hepatocellular carcinomas was significantly correlated with a low grade of differentiation and a shortened overall survival (P<0.05). No significant differences were found between the expression of Bif-1 and age, gender, tumor size, damage of capsule, expression of hepatitis B surface antigen (HBs-Ag) and portal venous invasion. Our data demonstrated that Bif-1 is frequently expressed in hepatocellular carcinoma. Overexpression of Bif-1 is a new independent prognostic marker, which is associated with poor differentiation as well as shortened overall survival.