Serum-free conditions for the long term growth and differentiation of neoplastic canine keratinocytes.

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Abstract
Long term cultures of canine keratinocytes have been established but culture conditions currently used require supplementation with fetal bovine serum (FBS). Unfortunately, FBS contains many non-defined components which may interfere with in vitro studies. This study describes the development of defined serum-free culture conditions for neoplastic canine keratinocytes grown submerged and at the air-liquid interface. Two commercially available serum-free media established for human epidermal cells failed to support canine keratinocyte growth. In contrast, a defined serum-free medium developed in our laboratory successfully supported proliferation of neoplastic canine keratinocytes for at least 40 passages. Cells showed a slower growth rate, but reached similar final densities and were morphologically identical to those cultured in FBS. Grown at the air-liquid interface, the cells reached the same degree of differentiation as in vivo stratified squamous epithelium and cultures grown in FBS. These results demonstrate that canine keratinocytes require different serum-free growth conditions than human cells. Neoplastic canine keratinocyte cultures, grown under serum-free culture conditions, provide an ideal in vitro system for comparative studies of keratinocyte biology and pathogenesis of various dermatoses.