Summary Cell proliferation of transitional cell bladder cancer (TCC) was determined by PCNA (proliferating cell nuclear antigen)/cyclin immunostaining in 178 TCCs and the results were related to established prognostic factors, progression and survival during a mean follow-up period of 10 years. The fraction of PCNA/cyclin positive nuclei was related to T-category (P= 0.008), papillary status, WHO grade, DNA ploidy, S phase fraction, M/V index (volume corrected mitotic index) and AgNORs (silver stained nucleolar organiser regions) (for all P<0.001). TCCs presenting with pelvic lymph node metastasis at diagnosis had a significantly higher growth fraction than the tumours confined to the bladder wall (P<0.001). The fraction of PCNA/cyclin positive nuclei predicted progression in T-, N-and M-categories (P<0.001). In Ta-Ti tumours high fraction of PCNA/cyclin positive nuclei predicted metastasis (P = 0.019). In survival analysis the fraction of PCNA/ cyclin positive nuclei predicted survival in the entire cohort (P< 0.001) and in Ta-TI tumours (P = 0.0005). In a multivariate survival analysis the fraction of PCNA/cyclin positive nuclei showed independent predictive value in the entire cohort (P = 0.046), in papillary tumours (P= 0.006) Wallace, 1985) and the treatment of patients has been detailed previously (Lipponen et al., 1990b(Lipponen et al., , 1991b UICC 1978 (UICC, 1978 and it was based on the above mentioned examinations added with the pathologists reports. The follow-up investigations were done at 3 month intervals during the first 2 years and thereafter at 6 months intervals. If a recurrent growth was observed the follow-up program was started again. The treatment of recurrent tumours was based on the same principle as the treatment of primary tumours. Many of the patients who died were autopsied to ascertain the extent and metastasis of tumours at the time of death.
Histological methodsThe histological samples were preoperative biopsy specimens. The samples were fixed in buffered formalin (pH 7.0), embedded in paraffin, sectioned at 5 gm and stained with hematoxylin and eosin or Van Gieson stains for grading. The samples were graded according to WHO (Mostofi, 1973). The distribution of cases into WHO grades and T-categories is shown in Table I. The papillary status of tumours was recorded and the tumours were divided into papillary (n = 149) and nodular (n = 29) types.
ImmunohistochemistryFor immunohistochemical demonstration of PCNA/cyclin, 5 flm sections from the primary TCCs were deparaffinised and rehydrated. Endogenous peroxide was blocked by 3% hydrogen peroxide for 5 min followed by a wash for 5 min with PBS. The tissue sections were incubated with the antiCorrespondence: P. Lipponen,