Human blood-borne monocytes were cultured for up to 22 days on disposable Teflon foils. Within 8 days, these monocytes developed into mature macrophages. At various stages of differentiation, the cells were recovered from the hydrophobic membrane and were assayed for typical monocyte-macrophage enzymes and morphology, binding of monoclonal antibodies (OKM1, OKla1), Fc and transferrin receptors, phagocytic activity, lysozyme production, and ability to inhibit the growth of an allogeneic tumor target cell line (U937). A significant antitumor activity of mature macrophages was found, which developed along with the differentiation of the monocyte precursor cells. In addition, cytotoxic effector macrophages could be activated by lymphokine-rich medium and synthetic alkyl-lysophospholipids. After density gradient separation, light cells (less than 1.05 and less than 1.06 g/ml) showed enhanced cytotoxicity, whereas cells from the dense fraction (greater than 1.06 g/ml) with low base-line activity could be best activated for cytotoxicity by lymphokines. If monocyte-macrophages are involved in a natural surveillance mechanism, our results may indicate the importance of unimpaired macrophage maturation to generate effective host defense against tumor development.