Nutritional and regulatory roles of human serum in cultures of human granulopoietic cells.

We proposed to develop a tissue culture medium supplement based on human serum that would support proliferation of human marrow cells and then to use it in an examination of human serum for factors that regulate colony formation. We examined various serum treatments ( heat, dialysis, ultrafiltration) and found that heat-treated human serum (58#{176}Cfor 120 mm) retained the ability to support proliferation and that this heat treatment eliminated the variability between different sera at concentrations 30%. We concluded that tissue culture medium, supplemented with 30% heat-treated human serum, was adequate and that the use of this growth medium would complement an examination of human serum for thermolabile factors. To reduce cellcell interactions in culture to a minimum, we used cell separation procedures and employed nonadherent light-density bone marrow cells (NALD BMC) for colonyforming cells, light-density white blood cells (LD WBC) for colony-stimulating cells, and LD WBC-conditioned medium for colony-stimulati ng factor. We examined human serum, whole and fractionated (Sephadex G200), for effects on colony formation in agar cultures supplemented with 30% heat-treated human serum contaming NALD BMC only, NALD BMC plus LD WBC, and NALD BMC plus conditioned medium. We found that human serum contamed thermolabile inhibitory and stimulatory activities that regulated colony formation in NALD BMC cultures stimulated with LD WBC. Neither whole nor fractionated human serum induced colony formation in NALD BMC cultures. This suggests that the colony-stimulating activity of human serum is not due to colony-stimulating factor.