The Eagle medium containing the epidermal growth factor (EGF), insulin, transferrin and a source of iron is able to support proliferation of 3T6 cells in the absence of serum. The formation of 3T6 cell clones in the similar medium supplemented with non-essential amino acids confirms the usefulness of such a medium for cell cultivation. It has been found that the exclusion of EGF rather than other above components from the medium inhibits most highly the increase in cell number.