The interaction of lactoferrin from human and bovine milk with the human promonocytic cell line U937 has been studied. Both human and bovine Fe-lactoferrins bound to the cells. Binding of bovine lactoferrin was inhibited by excess bovine lactoferrin but not by human lactoferrin, suggesting that the binding mechanisms for the two proteins are different. Binding of human but not bovine lactoferrin was inhibited by bovine lactoperoxidase, while a 20-fold excess of human IgA inhibited binding of human but not bovine lactoferrin. Human and bovine α-lactalbumins, bovine β-lactoglobulin, and human lysozyme had no effect on binding of lactoferrin from either species. Samples of bovine Fe- and apolactoferrin in capillary tubes were exposed to temperatures of 72 °C for 20 s, 85 °C for 20 min or 137 °C for 8 s. All the heated samples inhibited binding of native Fe- and apolactoferrin, though to a lesser extent than the native proteins. Both heated and native lactoferrins enhanced [3H]thymidine incorporation by U937 cells, except for Fe-lactoferrin heated at 85 °C for 20 min, which was inhibitory. These results suggest that heat treatment of lactoferrin under conditions used for industrial processing does not greatly affect its ability to interact with and stimulate monocytic cells, and that other milk proteins in general do not interfere with lactoferrin–monocyte interactions. It may thus be feasible to incorporate biologically active lactoferrin into infant formulas.