Abstract Purpose We performed this study to evaluate the role of endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) in the pathologic diagnosis of lung cancer including lung masses as well as lymph nodes as targets. Methods We retrospectively reviewed 126 patients who underwent EBUS-TBNA to diagnose radiologically suspected lung cancer. The patients had masses or lymph nodes that were highly suspicious for malignancy and accessible by EBUS-TBNA. Results EBUS-TBNA was performed on 195 lesions (lymph nodes, n = 151; lung masses, n = 44). In 61 cases, other diagnostic methods had failed previous to EBUS-TBNA. In 118 patients, no definite endobronchial mucosal tumor invasion was observed. In eight patients with endobronchial tumor invasion, EBUS-TBNA was chosen due to tumor bleeding, necrosis, or difficult location for endobronchial biopsy. EBUS-TBNA confirmed 105 lung cancers, five other malignancies and six specific benign cases, demonstrating a diagnostic yield of 92.1% (116/126). Nine cases were diagnosed by other methods (lung cancer, n = 2; other malignancies, n = 2; benign cases, n = 5). One case that was not confirmed by any diagnostic method was considered false negative. The sensitivity and diagnostic accuracy of EBUS-TBNA in the diagnosis of lung cancer were 97.2% (105/108) and 97.6% (123/126), respectively. Conclusions EBUS-TBNA targeting lymph nodes or masses highly suspicious for malignancy demonstrated high diagnostic value in the diagnosis of lung cancer. EBUS-TBNA is recommended for these cases, especially when other diagnostic methods have failed or are difficult.