Long-term production of hemopoietic progenitors in cultures containing low levels of serum.

A new low-serum-containing medium is described for long-term murine bone marrow cell cultures. This medium consists of a 1:1 mixture of an enriched Dulbecco's modified Eagle's medium (EMED) and a modified Ham's nutrient mixture F-12 (FMED) supplemented with fetal calf serum (1% vol/vol), bovine serum albumin (1 mg/ml), human transferrin (78 micrograms/ml), linoleic acid (2.1 micrograms/ml), cholesterol (2.6 micrograms/ml), hydrocortisone (10(-5) M), and alpha-thioglycerol (7.5 X 10(-5) M). Hemopoietic progenitor cell production was maintained for at least nine weeks in cultures established with a single inoculum of marrow cells and fed weekly by removal of all nonadherent cells and growth medium. The kinetics of hemopoiesis in these cultures was highly reproducible, facilitating quantitative comparison between experimental groups. Coating the flasks with serum proteins enhanced adherent layer formation and increased progenitor cell production compared to cultures established in untreated flasks. Hemopoietic cell production was, in fact, higher in this medium than in the mixture of EMED and FMED supplemented with 20% serum.