Growth and differentiation of human and murine erythroleukemia cell lines in serum-free synthetic medium.

Only two chemicals (transferrin and selenium dioxide) are required to supplement serum-free Roswell Park Memorial Institute Medium 1640 for long-term growth and for spontaneous and induced differentiation of established lines of human and mouse erythroleukemia cells. We describe here two serum-free media (a minimal synthetic medium and a high-density synthetic medium) that support the growth and differentiation of human K562(S) and mouse clones 745, 707, and 3TCl 12 erythroleukemia cell lines in long-term culture. The doubling times of the erythroleukemic cell populations are longer in minimal synthetic medium that in serum-containing medium. Cell saturation density in minimal growth medium is one-half that obtained in serum-containing medium for clone 745, whereas for K562(S) it is approximately the same. Cell saturation density in high-density medium (containing albumin) is greater than that achieved in serum-containing medium for K562(S), whereas for clone 745 cell saturation density increases for cells in midlogarithmic growth, although not to the density of cells grown in serum-containing medium. The differences in saturation density are due to a decreased doubling time as well as to better survival of the cells 3 or 4 days after plating. The cells can grow in the synthetic media and be passaged for as many generations as desired without impairment of growth capabilities. In the minimal synthetic medium, spontaneous differentiation of erythroleukemia cells continues to occur, indicating that spontaneously differentiating cells are the result of intracellular mechanisms controlling the expression of a genetic program of some of the cells at any given time. Hemoglobin synthesis can be induced in cells growing in synthetic medium by using lower concentrations of the same inducers that are effective in serum-containing medium, indicating that these chemicals do not depend on serum factors to initiate the process of differentiation. The percentage of benzidine-positive cells and the concentration of hemoglobin per cell, however, are less in the synthetic medium than in serum-containing medium, suggesting that serum factors do play a role in modulating the extent of hemoglobin synthesis. The types of hemoglobins synthesized by cells in synthetic medium are identical to those reported in serum-containing medium.