Objective: This study investigated whether liquid-based cytology (LBC) can improve diagnostic values of cytological assessment of endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA). Study Design: A cohort of 600 cases in West China Hospital was prospectively studied from June 2012 to September 2013. EBUS-TBNA was carried out in outpatients under local anesthesia and moderate sedation. The procedure was performed with an echobronchoscope (BF-UC160F-OL8, Olympus, Tokyo, Japan). Histological cores were stained with hematoxylin and eosin for further study. Additional immunohistological analysis was performed for establishing a reliable diagnosis when necessary. Aspirates were smeared on glass slides and separate aspirates were processed by the monolayer SurePath method. Results: In total, 480 malignant tumors and 120 benign lesions were confirmed by histological examination. The sensitivity of SurePath liquid-based preparations and conventional smears was 82.1 and 56%, and the specificity was 87.5 and 82.5%, respectively. The combined specificity was 100%. Positive predictive values of the two groups were 96.3 and 92.8%, whereas negative predictive values were 54.9 and 31.9%, respectively. The difference between the two groups was significant (p < 0.001). Conclusions: LBC preparation can improve cytological assessment of EBUS-TBNA. Histological study is necessary in cases in which the cytological diagnosis is obscure.