OBJECTIVES: The aim of this study was to investigate microRNAs related to postoperative prognosis of pancreatic cancer by applying microarray analysis to stored surgical specimens. METHODS: We retrospectively analyzed the relationship between microRNA expression and postoperative survival of pancreatic cancer patients in Samsung Medical Center. After the correlation of microRNA expression between paraffin-embedded and fresh-frozen specimens was confirmed, we applied NanoString nCounter system to formalin fixed paraffin-embedded surgical specimens between 1995 September and 2010 June. RESULTS: Paired fresh and paraffin-embedded samples from 9 patients showed a high correlation (correlation coefficient > 0.91). Among 734 microRNAs, 7 microRNAs were found to be related to postoperative overall survival of 221 pancreatic cancer patients: hsa-miR-99a, hsa-miR-150, hsa-miR-194, hsa-miR-375, hsa-miR-664, hsa-miR-342-3p, and hsa-miR-487b (P < 0.01). The prognostic microRNA signature was consistent after adjustment of clinical parameters: resection status, stages, age, sex, adjuvant treatment, pathological differentiation and postoperative carbohydrate antigen 19-9 levels. CONCLUSIONS: We found a prediction model for postoperative prognosis of pancreatic cancer using 7 microRNA expression patterns from archived formalin-fixed paraffin-embedded surgical specimens. The methods for microRNA detection and new microRNAs detected in this study provide new insights for new research of surgically resected pancreatic cancer samples and microRNAs.