We measured the concentrations of protein, insulin, and epidermal growth factor (EGF) in human milk from mothers delivering at term. Samples were obtained from d 1 (colostrum) to d 42 after birth. Human colostrum contains very high concentrations of protein [83.7 ± 7.4 g/1 (SEM)], insulin (3.75 ± 0.88 nM), and EGF (53.9 ± 6.9 nM). Similar concentrations have been measured in prebirth milk. Insulin, EGF, and protein in milk decline rapidly during the first few days of lactation but remain constant thereafter. Although the concentrations of insulin and EGF in mature milk are only 10% of those in colostrum, they are considerably higher than in serum. We also showed that human milk has a growth-promoting activity in cultured cells, causing a stimulation of protein synthesis in L6 myoblasts and 3T3-L1 fibroblasts and an increase in DNA synthesis in L6 cells and T47D breast cancer cells. This mitogenic activity declines as lactation progresses, with a similar time-course to the fall in insulin and EGF; however, the cell lines used here are not responsive to either of these two growth factors in the range of concentrations found in milk. This indicates that human milk also contains high concentrations of additional, unidentified growth factors. The occurrence of high concentrations of growth factors in milk suggests that they may be important for the proliferation and differentiation of infant tissues.