Background: Preoperative differentiation of benign and malignant/potentially malignant pancreatic cystic lesions is problematic. Data to support the role of EUS and EUS-guided fine-needle aspiration (EUS-FNA) are limited. This study assessed the sensitivity, specificity, and accuracy of EUS, cytopathology, and analysis of cyst fluid for pancreatic cystic lesions. Methods: Retrospectively, 111 consecutive patients were identified (54 men, 57 women; mean age 59 years, range 18-79 years) who underwent EUS from July 1997 to September 2000 because of known or suspected pancreatic cystic lesions based on CT or transabdominal US. Thirty-four patients (16 men, 18 women; mean age 55 years, 25-79 years) who underwent surgery formed the basis for this analysis. EUS diagnosis was compared with surgical pathology. Selected patients underwent EUS-FNA to obtain specimens for cytopathologic analysis and for determination of carcinoembryonic antigen levels. Based on surgical pathology, cysts were classified as benign (simple cyst, pseudocyst, serous cystadenoma) or malignant/potentially malignant (mucinous cystadenoma, intraductal papillary mucinous tumor, cystic islet cell tumor, cystic adenocarcinoma). Results: EUS-FNA with cytopathologic assessment of cyst fluid was performed for 18 of the 34 patients; carcinoembryonic antigen level was determined in 11 cases. For EUS, cytopathology, and carcinoembryonic antigen, sensitivity was, respectively, 91%, (p = 0.01 vs. cytology), 27%, and 28%; specificity was, respectively, 60%, 100%, and 25%; and, accuracy was, respectively, 82%, 55%, and 27%. The sensitivity of EUS in all 13 patients with cystic islet cell tumor, intraductal papillary mucinous tumor, or cystic adenocarcinoma was 100%. Combining EUS, cytopathology, and carcinoembryonic antigen results did not improve accuracy. There were no complications related to the EUS or EUS-FNA. Conclusions: EUS alone is sensitive and accurate in identifying malignant/potentially malignant pancreatic cystic lesions. EUS-FNA to obtain specimens for cytopathologic analysis and determination of carcinoembryonic antigen levels, although safe, does not enhance diagnostic yield. (Gastrointest Endosc 2002;56:543-7.)