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Objective To investigate the diagnostic value of chest CT scan combined with telomerase activity and p16 gene methylation from exfoliated cells of sputum in 55 cases of solitary pulmonary nodules (SPN; ≤30 mm)suspected early peripheral lung cancer. Methods The sputum specimens from 34 cases of cancer nodules and 21 cases of benign lesion were detected for telomerase activity by TRAP-PCR-ELISA and p16 gene methylation by PCR-based methylation analysis. Results The qualitative diagnostic accuracy of CT scan was 61.8%(34/55) for SPN provided by pathology. Cytology analysis of sputum was positive in 13 cases (38.2%). Telomerase activity was positive in 29 cases: sensitivity was 79.4%, specificity was 90.5%, accuracy was 83.6%; p16 gene methylation was found in 11 cases: sensitivity was 32.4%, specificity was 100.0%, and accuracy was 58.2%. The sensitivity was increased to 86.1% by combination of telomerase activity and p16 gene methylation. Compared with nodules without malignant CT signs, expression of telomerase activity and p16 methylation of SPN with malignant CT signs (lobulation or spiculate protuberance or spicule sign) had a significant difference (P<0.01). Conclusion The results suggest that chest CT scan combined with telomerase activity and p16 gene methylation detection in sputum for patients with peripheral lung cancer may enhance the diagnostic value of radiology and conventional cytology.