Building a protocol for culturing normal human epidermal Keratinocytes

Abstract

Human epidermal keratinocytes (HEKs) are the most common cell type in the epidermis. Most of 90% of cells in epidermis are keratinocytes. Although the methods for keratinocyte culture from epidermis have been successfully in laboratory and clinical usage, building an optimal culture protocol that support their high proliferative capacities and adhesion ability in vitro is a primary factor interfering with the efficiency in their usage. This research is conduced to identify the optimal procedure for representing a high attachment and growth rates of HEKs in free-serum environment. Firstly, the human skin fragments are proteolytically digested by 0.25% trypsin/0.02% EDTA. Keratinocytes are obtained from the epidermis and cultured. The cell survival rate was determined by dye exclusion (trypan blue). In conclusion, this HEKs culture protocol with serum free medium resulted in the highest survival rate (82.95%) and also in the highest yields of separated cells (6.8x 106cells/cm2). These results demonstrate that this defined culture model has a potential application in clinical usage and can be used for further related research.

Keywords: Human Epidermal Keratinocytes, HEKs, Free-Serum Medium, Trypsin/EDTA, Trypan blue