We have adapted a human monocytic leukemia cell line [THP-1c12(+)] so that it can proliferate in a completely protein-free chemically defined medium of an equal mixture of Dulbecco's modified Eagle's minimum essential medium and Ham's F12. This cell line was designated as THP-1c12(-). When a sufficient number of THP-1c12(-) cells was seeded in culture, the maximum cell density after 6 days of culture was 1 X 10(6)/ml with a doubling time of 30 hr. Transferrin showed a slight stimulative effect on the growth of THP-1c12(-) cells, but insulin did not. The surface profile of THP-1c12(-) was the same as that of THP-1c12(+), which possessed Mol. Mo5, LeuM3, My9 and Ia-like antigens as well as a large number of Fc receptors. Phagocytic ability with respect to IgG-coated sheep red blood cells was evident in both THP-1c12(+) and THP-1c12(-) cells. A small percentage of THP-1c12(-) cells retained the ability to reduce nitroblue tetrazolium when stimulated with 12-tetradecanoyl-phorbol-13-acetate. Culture supernatant from THP-1c12(-) stimulated the incorporation of 3H-thymidine into its own cells, suggesting an autocrine mechanism by which the growth of THP-1c12(-) cells is induced in a protein-free medium.