The archival paraffin-embedded specimens from 63 ampulla of Vater cancers after pancreaticoduodenectomy between 1965 and 1991 were analyzed by flow cytometry. Of the 63 cancers, 31 (49.2%) were diploid DNA cancers and 32 (50.8%) were aneuploid. Patients with diploid DNA cancer had a median survival time of 159.0 months, and patients with aneuploid DNA cancer had 24.0 months. This difference is statistically significant (P = 0.0257). The aneuploid group did have a poorer prognosis than the diploid group. The multivariate analysis demonstrated that DNA ploidy was an independent and very important prognostic factor, even stronger than the stage and lymph node status. There was a tendency toward higher values of S-phase fraction, proliferative index, and total aneuploid DNA fraction in the shorter survival groups, but they were of no statistical significance. These data suggest that DNA ploidy appears to be the most important and the only pre-operative predictor of prognosis in resectable ampulla of Vater cancers since endoscopic biopsy is feasible.