Primary cultures of cells derived from 13 patients with acute myelomonocytic leukemia (AMML) were studied with particular emphasis on in vitro proliferation, cell differentiation and the mode for establishment of cell lines. Using irradiated human macrophage monolayers to assist cell growth, we obtained four new cell lines of myelomonocytic origin. All the cell lines were characterized for cytochemical markers and response to phorbol esters (TPA), a differentiation inducing agent. In the absence of any inducing agent, spontaneous differentiation of blast cells into mature macrophages-like cells occurred in 8 out of the 13 primary cultures. Thus, maturation induction by agents such as TPA is not always required in order to obtain leukemic cell differentiation in vitro. The regulation of cell proliferation and differentiation by cellular interactions and by extrinsic soluble products is discussed in detail, in the light of these findings.