BACKGROUND & AIMS: Despite the widespread use of endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) to sample pancreatic lesions and the standardization of pancreaticobiliary cytopathologic nomenclature, there are few data on inter-observer agreement among cytopathologists evaluating pancreatic cytologic specimens obtained by EUS-FNA. We developed a scoring system to assess agreement among cytopathologists in overall diagnosis and quantitative and qualitative parameters, and evaluated factors associated with agreement.; METHODS: We performed a prospective study to validate results from our pilot study that demonstrated moderate to substantial inter-observer agreement among cytopathologists for the final cytologic diagnosis. In the first phase, 3 cytopathologists refined criteria for assessment of quantity and quality measures. During phase 2, EUS-FNA specimens of solid pancreatic lesions from46 patients were evaluated by 11 cytopathologists at 5 tertiary care centers using a standardized scoring tool. Individual quantitative and qualitative measures were scored and an overall cytologic diagnosis was determined. Clinical and EUS parameters were assessed as predictors of unanimous agreement. Inter-observer agreement (IOA) was calculated using multi-rater kappa (kappa) statistics and a logistic regression model was created to identify factors associated with unanimous agreement.; RESULTS: The IOA for final diagnoses, based on cytologic analysis, was moderate (kappa = 0.56; 95% CI, 0.43-0.70). Kappa values did not increase when categories of suspicious for malignancy, malignant, and neoplasm were combined. IOA was slight to moderate for individual quantitative (kappa = 0.007; 95% CI, -0.03 to -0.04) and qualitative parameters (kappa = 0.5; 95% CI, 0.47-0.53). Jaundice was the only factor associated with agreement among all cytopathologists on multivariate analysis (odds ratio for unanimous agreement, 5.3; 95% CI, 1.1-26.89).; CONCLUSIONS: There is a suboptimal level of agreement among cytopathologists in the diagnosis of malignancy based on analysis of EUS-FNA specimens obtained from solid pancreatic masses. Strategies are needed to refine the cytologic criteria for diagnosis of malignancy and enhance tissue acquisition techniques to improve diagnostic reproducibility among cytopathologists.