[Culturing of kerationocytes in serum-free medium]

ABSTRACT

To establish a simple method for preparation of pure epidermal cell in serum-free medium, without co culturing with lethally irradiated 3T3 cells. A piece of skin biopsy was taken from a rat. After trypsinazation to separate epidermal layer, the epidermis was cut into explants 1x1 mm and were laid at 50 mL on tissue cultured flasks. Then, the explants were covered with serum-free keratinocyte growth medium, while the culture medium was changed every two days. The keratinocyte colonies were expanded in flasks and proliferated to 70% confluency on day 21. After two subcultures, the cells were frozen. Pure epidermal cells are established by this technique in a serum-free keratinocyte growth medium, under feeder cell-free condition. Culturing of keratinocytes in serum-free medium is proved to be a useful and simple method for keratinocyte isolation. The successful culturing of keratinocytes from a small skin biopsy can be useful in the treatment of major burn wounds