Objective: To investigate the expression of paired-related homeobox 1 protein (Prrx-1) in gallbladder cancer tissue and its significance. Methods: The Prrx-1 expressions in 45 specimens of gallbladder cancer tissues and 35 specimens of gallbladder tissues with benign diseases were determined by immunohistochemical staining, and the relations of Prrx-1 expression with clinicopathologic profiles of gallbladder cancer were analyzed. Results: The expression rate of Prrx-1 was 55.6% (25/45) in gallbladder cancer tissue, and was 5.7% (2/35) in gallbladder tissue with benign diseases, and the difference between them had statistical significance (χ2=21.87, P<0.05). Statistical analysis indicated that the Prrx-1 expression was closely associated with the degree of differentiation and clinical stage of gallbladder cancer (both P<0.05), but irrelevant to the sex and age of the patients and histological type of gallbladder cancer (all P>0.05). Conclusion: Prrx-1 expression is increased in gallbladder cancer and its expression may be closely connected to the occurrence and development of gallbladder cancer.