Objective: To detect the expression of Rab27b in human colorectal cancer (CRC) and investigate the relationship between Rab27b expression and clinical characteristics, especially the prognosis of CRC. Methods: Immunohistochemistry analysis by tissue microarrays (80 CRC samples and 20 tumor-adjacent tissue samples) was performed to characterize Rab27b expression in CRC. Cox regression and Kaplan-Meier survival analyses were carried out to evaluate the prognosis of CRC. Results: Immunohistochemistry analysis showed that the expression of Rab27b in CRC was significantly higher than that in normal tumor-adjacent tissues (P = 0.041). Moreover, the expression level of Rab27b protein in CRC was significantly related to gender (P = 0.012), serum CEA level (P = 0.014), distant metastasis (P = 0.021) and TNM stage (P = 0.013). COX multi-factor analysis showed that Rab27b expression (P = 0.021) could be recognized as an independent prognosis factor for CRC. Conclusion: Rab27b expression was abnormally highly expressed and correlated with malignant phenotypes of CRC. It may serve as a novel prognostic factor for CRC.