Objective: To investigate the relationship between the expression of EphB4 and HIF-1α in lung cancer and their biological significance. Methods: The expression of EphB4 and HI F-1α was detected in 54 specimens of lung cancer by using streptavidin-peroxidase (SP)immunohistochemical method, and 10 normal lung tissues as control. Results: Among the 54cases of lung cancer, the positive rate of EphB4 was 50.0%; 42.6% of the tumors were positive for HIF-1α. The expression of these two kinds of proteins was related to gross types, differentiation and clinical stages (P＜0.05), but not to histological classification, age, sex and lymphoid metastasis (P＞0.05). A highly positive correlation was observed between the expression of EphB4 and HIF-1α (P＜0.01). Conclusion: Overexpression of EphB4 and HIF-1α may play an important role in the pathogenesis, progression and malignant degree of lung cancer. Detection of EphB4 and HIF-1α expression might be helpful to predict prognosis of patients with lung cancer.