Multiparameter flow cytometry studies were performed on clinical samples of human bladder tumors to simultaneously analyse DNA content and the expression of surface glycoproteins defined by monoclonal antibodies T16, Om5, T43, and T138. The results of tests performed on 80 samples of bladder irrigations and tumors from 68 patients were correlated with clinical findings at the time of sampling and with disease outcome prospectively (mean follow-up 2 years). Measuring the level of the panurothelial antigen T16 provided more precision in DNA analysis and served as an internal standard to measure the relative expression of the other cell surface antigens studied. The panel of monoclonal antibodies improved the analytical capacity to study the heterogeneity of antigenic phenotypes within individual samples. Aneuploidy frequently correlated with high stage cancers and with a high rate of clinical cancer progression defined as metastasis or death by cancer. However ploidy was not an entirely reliable prognostic indicator since a significant proportion of Ta and T1 nonprogressing tumors were aneuploid, while in 6/20 cases of cancer progression, the samples were near diploid. Contrary to Om5, T43 and T138 antigens were expressed significantly more often on aneuploid samples, although they appear to provide additional information. T138 was positive on 17/18 samples from patients with high stage cancers of which five were near diploid. It was also positive on 4/5 samples from patients with Ta and T1 tumors in whom disease progressed to metastasis and death. Overall, the expression of T138 antigen was a better single indicator of clinical cancer progression than was ploidy. Further stratification was obtained with combined results of DNA and T138 antigen studies. Within the near diploid group, the incidence of bladder cancer death was 0/26 for T138 negative and 5/13 (38%) for T138-positive patients (P less than 0.01). In the aneuploid group incidence of bladder cancer death was 2/10 (20%) for T138-negative and 12/19 (63%) for T138-positive patients (P less than 0.05). These results suggest that simultaneous flow cytometry measurements of DNA and surface antigens may better assess the prognostic behavior of human bladder tumors.