AbstractWe have examined conditioned medium (CM) from cultures of normal rat mammary epithelial (RME) cells for growth factor activity on fresh RME cell cultures. RME cell‐derived CM contained potent growth inhibitory activity toward fresh RME cell cultures when the medium was acidified by dialysis against 1% acetic acid prior to concentration. Dialysis of the CM at neutral pH resulted in CM that had growth stimulatory activity and no inhibitory activity. The acid‐activated growth inhibitor was heat and acid stable, protease sensitive, and eluted from a Bio‐Gel p60 column with a peak of activity in the 28 kDa range. Incubation of the acidified‐concentrated CM with neutralizing antiserum (affinity purified IgG) against transforming growth factor (TGF)‐β completely abolished the inhibitory activity of the CM. Furthermore, RME cell growth in the presence of the growth inhibitor plus TGF‐β antiserum was greater than that observed in growth medium alone. Subsequent experiments demonstrated that addition of TGF‐β antiserum alone to serum‐free medium enhanced RME cell growth, whereas addition of nonimmune IgG was without effect even at 25‐fold higher concentrations. Zymographic analysis of RME‐CM revealed the presence of plasminogen activator proteases that may mediate the partial activation of the latent growth factor. These results indicate that normal RME cells secrete a latent TGF‐β‐like growth factor into conditioned medium. Furthermore, the results indicate that some of the latent growth factor is activatedin situand contributes to the growth potential of the cells in primary cul