A431 human epidermoid carcinoma cells in culture may be grown in the absence of serum in a one to one mixture of Ham's F12 and Dulbecco-modified Eagle's medium supplemented with 10 micrograms/ml bovine insulin, 10 micrograms/ml human transferrin, 5 micrograms/ml human cold-insoluble globulin and 0.5 mM ethanolamine. Growth rate of the cells in this serum-free medium is essentially identical to that of cells in medium containing 10% fetal calf serum. Addition of epidermal growth factor at concentrations which are stimulatory for the growth of other cell types in culture causes and inhibition of A431 cell growth in both serum-free and serum-containing medium. The inhibitory effect of epidermal growth factor is reversible upon replacement of epidermal growth factor-containing medium with medium containing no epidermal growth factor. Simultaneous addition to the medium of antibody to epidermal growth factor along with epidermal growth factor prevents the inhibitory effect.