The interpretation of pancreas fine-needle aspiration (FNA) is extremely difficult given the cytologic overlap of neoplastic and reactive processes. Using serial analysis of gene expression, we have discovered 2 new markers of pancreatic adenocarcinoma, mesothelin and prostate stem cell antigen (PSCA), and confirmed their specificity by immunohistochemical labeling. Here we evaluate the potential contribution of immunohistochemical labeling of mesothelin and PSCA to the interpretation of pancreas FNAs. Thirty pancreas FNAs with follow-up data were reviewed. Unstained cell block sections from these aspirates labeled for mesothelin and PSCA using immunohistochemistry were compared with initial cytologic diagnoses and with follow-up diagnoses. On follow-up, 19 patients proved to have cancer, and 11 did not. Initial cytologic diagnosis of malignancy correlated with carcinoma on follow-up in 12 of 12 cases, and initial benign cytologic diagnosis correlated with benign follow-up in 8 of 9 cases (sensitivity, 92%; specificity, 100%). Six of the 9 patients with suspicious cytology were found to have a carcinoma on follow-up. PSCA labeling was present in 16 of the 19 patients who ultimately were proven to have carcinoma; PSCA labeling was absent in 10 of the 11 lesions proven to be benign (sensitivity, 84%; specificity, 91%). Mesothelin labeling was present in 13 of the 19 patients who ultimately were proven to have carcinoma; mesothelin labeling was absent in 10 of the 11 lesions proven to be benign (sensitivity, 68%; specificity, 91%). Five of the 6 cytologically suspicious cases with malignant follow-up labeled for either PSCA or mesothelin (83%), and 2 of the 6 cases labeled for both markers. None of the 3 suspicious cases with benign follow-up labeled for either PSCA or mesothelin. Increasingly, molecular techniques are identifying potential cancer markers that may have diagnostic utility. In this study, immunohistochemical labeling for 2 of these markers, PSCA and mesothelin, appears highly specific for pancreatic adenocarcinoma in FNA specimens and useful in categorizing cytologically suspicious lesions.