ISOLATION AND LONG-TERM CULTURE OF HUMAN HAIR-FOLLICLE MELANOCYTES

We report a method to establish long-term cultures of melanocytes deri ved from human hair follicles. Normal human scalp was transected 1 mm below the epidermis, and hair follicles in the remaining dermis were i solated by collagenase treatment. Hair-follicle cell suspensions were prepared by trypsin/ethylenediamine tetraacetic acid treatment and cul tured in a mixture of Eagle's minimum essential medium (supplemented w ith 12-O-tetradecanoyl-phorbol-13-acetate and cholera toxin) and kerat inocyte serum-free medium. After contaminating fibroblasts and keratin ocytes were removed, cells with two distinct morphologies remained. Th ese included large, dendritic and deeply pigmented cells, which did no t proliferate and which disappeared by the third passage, and small bi polar cells, which initially were unpigmented, proliferated very rapid ly, and became pigmented after the addition of 3-isobutyl-1-methylxant hine to the culture medium. Both cell types were melanocytes as confir med by electron microscopy and by staining with antibodies to S-100, G (D3), and melanosomal antigens. The availability of cultured hair-foll icle melanocytes will facilitate investigations of the role of these c ells in normal and abnormal hair biology.