In order to assess the response of acute myeloid leukemia colony-forming cells (AML-CFU) to recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF), AML blasts of 20 patients were cultured in a colony assay supplemented with titrated concentrations of rGM-CSF. In 16 cases rGM-CSF was able to induce AML colonies. In eight cases maximal clonogenic cell proliferation was obtained with 100 U rGM-CSF/ml alone (type I response). In eight other cases, however, maximal colony numbers were reached only after the addition of low concentrations of PHA-leukocyte conditioned media (PHA-LCM) to the rGM-CSF containing cultures (type II response). These values could not be obtained with higher doses of rGM-CSF (500 U/ml) or PHA-LCM separately. Thus in this subgroup, optimal AML colony formation depended on rGM-CSF plus an additional factor. Finally, in 4 of 20 cases rGM-CSF alone (100 U, 1000 U/ml) was not capable of inducing any AML colonies in vitro (type III). In these latter cases proliferation of AML-CFU could be achieved only by supplementing PHA-LCM. We conclude that GM-CSF is a stimulator of the in vitro proliferation of AML clonogenic cells. However, in a majority of these cases, i.e., 12 out of 20, AML-CFU require an additional factor for optimal proliferation which is produced by PHA-stimulated leukocytes.