The aim of this study was to reexamine the prognostic role of tumor cell kinetics measured by S-phase fraction (SPF) and to establish its clinically relevant threshold values. SPF was determined by flow cytometry in a group of 920 consecutive breast cancer patients, all followed at our institute for 10 years (1988 to 1998). Mean age was 60.5 years (27-89 years). Median follow-up was 63 months (3-150 months). All patients had initial surgical treatment. SPF quartiles were: Q1=3.08%, median value = 5.98%, Q3=10.22%. A significant difference in overall specific survival was obtained between two populations divided by a cutoff at Q1 (p < 0.0001). A multifactorial analysis including SPF and known prognostic factors such as tumor size, node status, histological grade, ER and PR status was performed using the Cox model in a population of 719 patients: univariate analysis showed that each of these factors had significant influence on overall survival. Multivariate analysis selected three of them, ranked by decreasing order of hazard ratio (HR) value: SPF (HR: 3.88, p < 0.001), tumor size (HR: 2.49, p < 0.001) and nodal status (HR: 2.28, p < 0.001). In addition, when tumors were stratified according to SPF quartile values, there were statistically different overall survival curves in patients with small tumors (< 2 cm) and in axillary node-negative patients.