Preparation of Primary Cultures of Mouse Epidermal Keratinocytes and the Measurement of Phospholipase D Activity

Abstract

In this chapter information is provided about the outer layer of the skin, the epidermis, and the predominant cells comprising this epithelium, the keratinocytes. The evidence supporting a possible role for the lipid-metabolizing enzyme phospholipase D in regulating keratinocyte differentiation is also discussed. A detailed protocol for the preparation of primary cultures of epidermal keratinocytes from neonatal mice is described, to allow other investigators to obtain data concerning these important cells involved in forming and maintaining the mechanical and water permeability of the skin. Finally, a complete protocol for monitoring phospholipase D activity in intact cells is supplied in the hope that additional research will result in a better understanding of the role of phospholipase D in controlling keratinocyte proliferation and differentiation.