The requirements for the serum-free culture of PC13 murine embryonal carcinoma cells were determined. Supplementation of a 50:50 mixture of Dulbecco's modified Eagles medium and MCDB104 with transferrin (5 micrograms/ml), human high-density lipoprotein (HDL) (100 micrograms/ml), and human low-density lipoprotein (LDL) (50 micrograms/ml) supported growth comparable to that observed with 5% foetal calf serum. Media supplementation with lipoproteins apparently substitutes for the effects of insulin, desoctapeptide insulin, (DOP), or multiplication-stimulating activity (MSA) on EC cell multiplication. Clonal growth of PC13 EC cells in this serum-free medium could only be achieved in the presence of suitable feeder cell monolayers. These observations demonstrate that PC13 EC cells do not have an absolute requirement for exogenous mitogens to support multiplication.