The growth rate of DU145 prostate cancer cells in vitro is slowed considerably by changing the growth medium every 24 h, suggesting dependence upon endogenously-secreted growth factors. Because previous studies have identified epidermal growth factor (EGF) in the conditioned medium from DU145 cells, [35S]labeled EGF was selectively immunoprecipitated from the culture medium at 24-h intervals for quantitation. Under the culture conditions used, there was an initial phase of slow growth, the EGF level secreted per cell was highest on day 3 after plating, and an increase in cell number was most evident between days 3 and 4. Finally, growth was assayed under culture conditions where the medium was replaced every 24 h with fresh medium in the absence or presence of 10 ng/ml added EGF. The EGF was able to increase the cell growth up to the levels seen in cultures where the medium was unchanged during the entire period. We interpret these results as evidence that endogenously secreted EGF-like growth factors participate in an autocrine growth stimulation of DU145 cells.