High concentrations of fetal bovine serum induced colony formation in soft agar by anchorage-dependent, nontransformed mouse AKR-2B and rat NRK cells. The colony-stimulating activity in fetal bovine serum was precipitated by 45% saturated ammonium sulfate and migrated in molecular sieve chromatography as a single peak of activity in the 10,000-15,000 molecular weight range. The colony-stimulating activity was heat and acid stable and was destroyed by trypsin and dithiothreitol, indicating the activity is due to a polypeptide that requires disulfide bonds for biologicalactivity. No competition for binding to the epidermal growth factor receptor was associated with the colonystimulating activity. Isoelectric focusing revealed activity in the pI 4-5 range. The colony-stimulating activity in serum appeared to be of platelet origin because platelet-poor plasma and plateletpoorplasma-derived serum containedlittle activity, whereas acid/ ethanol extracts ofbovine and human platelets had potent colonystimulating activity. Chromatography of platelet extracts on BioGel P-60 revealed peaks of AKR-2B colony-stimulating activity in the 12,000 and 20,000 molecular weight ranges. The other biological and chemical properties of the platelet colony-stimulating activity were the same as those for the serum activity. The data indicate the presence in serum of a platelet-derived growth factor(s) with properties similar to those of the transforming growth factors.Cells that are normally anchorage dependent can be stimulated to form colonies when suspended in semisolid medium in the presence of higher than usual concentrations of serum. O'Neill et al. (1) reported that anchorage-dependent NIL-8 hamster fibroblasts exhibited a serum concentration-dependent growth in soft agarose and that the rate of growth of attached and suspended cells was approximately the same when the concentration of fetal bovine serum was raised to 66%. Peehl and Stanbridge (2) found that normal human fibroblasts would grow suspended in methylcellulose in the presence of 20% fetal bovine serum but not in medium with 10% fetal bovine serum; this anchorage-independent growth was enhanced by the addition of hydrocortisone.