The aim of the present study is to explore the role of annexin II in the development and progression of human non-small cell lung cancer (NSCLC). Real-time quantitative reverse transcription-polymerase chain reaction was conducted to detect annexin II mRNA expression. Annexin II protein expression was also determined by western blot. In addition, annexin II expression was analyzed by immunohistochemistry in 137 clinicopathologically characterized NSCLC cases. The correlation of annexin II expression with patients' survival rate was assessed by Kaplan-Meier analysis and Cox regression. Our results showed that the expression levels of annexin II mRNA and protein in NSCLC tissues were significantly higher than those in non-cancerous tissues. Immunohistochemistry analysis showed that annexin II expression was significantly correlated with tumor diameter, pathological grade, pT status, pN status, and pleural invasion. The results of the Kaplan-Meier analysis indicated that a high expression level of annexin II resulted in a significantly poor prognosis of NSCLC patients. Multi-variate Cox regression analysis revealed that annexin II expression level was an independent prognostic parameter for the overall survival rate of NSCLC patients. In conclusion, these results suggested that annexin II up-regulation was associated with poor prognosis in NSCLC; therefore, it might act as a prognostic marker and a new potential target for NSCLC treatment.