Medium 254 is a sterile, liquid tissue culture medium prepared with 200 μM calcium chloride for the culture of normal human epidermal melanocytes. This medium is intended for use as one component in a complete culture environment. When supplemented with HMGS or HMGS-2, Medium 254 will support the plating and proliferation of melanocytes at varying culture densities from 5 x 10³ cells/cm² to 1 x 10⁵ cells/cm². Additional applications for use may include primary isolation of melanocytes from skin.

Medium 254 is a basal medium containing essential and non-essential amino acids, vitamins, other organic compounds, trace minerals, and inorganic salts. This medium does not contain antibiotics, antimycotics, hormones, growth factors, or proteins. This medium is HEPES and bicarbonate buffered and designed for use in an incubator with an atmosphere of 5% CO²/95% air. To support plating and long-term proliferation of normal human melanocytes, Medium 254 must be supplemented with Human Melanocyte Growth Supplement (HMGS) or PMA-Free Human Melanocyte Growth Supplement-2 (HMGS-2). Each of these supplements contains all of the growth factors, hormones, and tissue extracts necessary for growth of melanocytes in Medium 254.