HMB‐45 monoclonal antibody recognizes an inducible and reversible melanocyte cytoplasmic protein

HMB‐45 is a monoclonal antibody directed against human melanoma cells and which stains epidermal and dermal melanoma cells, the junctional components of common and dysplastic mel‐anocytic nevi, and melanocytes in fetal skin. In addition, melan‐ocytes in a variety of reactive conditions have been shown to label with HMB‐45, as have dermal melanocytes within Spitz and dysplastic nevi. No melanocytes in normal adult epidermis or in the dermis of common nevi have stained with HMB‐45. In order to better understand the properties of this antibody, and of the melanocytes that react with it, we stained cultured human melanocytes grown in a variety of conditions. Melanocytes from human foreskins were grown for 2–3 weeks in MCDB 153 medium supplemented with insulin, epidermal growth factor, and bovine pituitary extract as a mixed population of keratinocytes and melanocytes. Some cells were transferred to basal medium MCDB 153 (unsupplemented) for periods ranging from 3–5 days, and a subset of these were returned to growth‐factor supplemented medium. In all cases, SI00 staining was used to confirm the presence of melanocytes. Melanocytes grown in complete medium showed strong granular cytoplasmic staining with HMB‐45. Cells transferred to basal medium showed a markedly diminished staining intensity which was reversible within 3 days upon return of the cells to complete medium. The findings suggest that expression of the protein recognized by HMB‐45 may be related to a growth factor present in complete medium, but missing from basal MCDB 153. Additional studies, supplementing medium with phorbol esters, markedly enhanced HMB‐45 staining and essentially removed keratinocytes from cell cultures. As determined by measurement of melanocyte proliferation, staining of HMB‐45 did not correlate with mitogenic activation but may correlate with biochemical activation by specific growth factors.