Objective To investigate the diagnosis value of CT-guided percutaneous transthoracic needle biopsy (TNB) combined with serum tumor markers for lung cancer. Methods The clinical data of the lung cancer patients treated in our hospital from January 2010 to January 2016 were analyzed. After inclusion and exclusion, 78 patients were enrolled in the study. Patients were divided into benign group (n=23) and malignant group (n=55) according to the TNB results. The TNB results, pathological results and complications of all patients were collected. Thereafter, the sensitivity, specificity and accuracy of TNB were analyzed. Also, the levels of CEA, NSE, and CYFRA21-1 were measured before and 12 months after surgery. Results 83.64% of malignant lung cancer patients were consisted of 65.22% of adenocarcinoma, 15.57% of squamous cell carcinoma and 15.22% of small cell carcinoma. 5.45% of non-malignant lung cancer patients included 66.67% of adenosarcoma, 33.33% of sarcomatoid carcinoma and 33.33%of malignant lymphoma. In the benign group, 60.87% of pneumonia patients included 57.14% of fungal pneumonia, 21.43% of mechanized pneumonia, 7.14% of granulation pneumonia, 13.04% of tuberculosis, 1.82% of pneumoconiosis, 8.70% of inflammatory pseudotumor, and 1.82% of fibrous tissue. According to the pathological results, malignant lung cancers and benign lung cancers accounted for 74.35% and 25.64%, respectively. The accuracy rate of pathological diagnosis in the malignant group was higher than that of the benign group (P<0.05). The sensitivity, specificity and accuracy of TNB in the diagnosis of lung cancer were 96.36%, 100.00% and 97.44%, respectively. The levels of CEA, NSE and CYFRA21-1 in the malignant group were significantly higher than those in the benign group before treatment (P<0.05), which were decreased in both groups after treatment, and were still lower in the benign group than in the malignant group (P<0.05). The complications including pneumothorax (2.56%), microbleeds (67.69%), chest tightness (12.82%) were cured after symptomatic treatment. Conclusion CT-guided percutaneous transthoracic needle biopsy combined with serum tumor markers is effective and safe to improve the detection rate of lung cancer, which can be used as an effective clinical diagnosis method.