Mitogen requirements of normal epidermal human melanocytes in a serum and tumor promoter free medium

Normal human melanocytes were cultivated in a serum free medium and the proliferation rate and antigen expression in the absence of each supplement was assessed every 3 days for 3 weeks. Results showed that the proliferation rate was significantly decreased in the absence of basic fibroblast growth factor, insulin and bovine pituitary extract as assessed 3 and 6 days of incubation (< 0.01). From day 9 through day 21, absence of transferrin, hydrocortisone, calcium, cholera toxin and epidermal growth factor was associated with a significant decrease of proliferation (< 0.001). Cells incubated with plain medium were almost absent from culture plates on day 6 and afterwards (< 0.0001). No evidence of contamination by epidermal and dermal cells was detected as all cultivated cells were labeled with MAbs HMB-45 and K.1.2.58. Absence of each supplement did not substantially affect antigen expression. It is concluded that basic fibroblast growth factor, insulin, and bovine pituitary extract are significant mitogens for melanocytes grown in a serum free medium from the very early phases of their growth.