The Isolation, Culture And Identification Of Keratinocyte And Fibroblast

Objective: To explose the cultivating technique of human keratinocyte and fibroblast under laboratory conditions, and to study the existence of epidermal stem cell in cultured human keratinocyte in vitro, and to explose the localization of CK19-positive-cells in various anatomic site of human skin. Methods: celt engineering thchniques including isolation, culture and subculture of human keratinocytes and fibroblasts were used, and to observe the pattern and growing course of cells.(l) By means of antibody of cytokeratin, the keratinocytes were identificated by immunohistochemistry. The cultured cells were analysed with the monoclone antibody of cytokeratin 19. (2) By means of antibody of viment, the fibroblasts were identificated by immunohistochemistry. (3) By means of the monoclone antibody of cytokeratin 19, to observe the localization of CK19-positive-cells in various anatomic sites of human skin. Result: 1 ,The epidermis were separated from the dermis intactly. The cultured cells were confirmed by means of electron microscopy and SP immunohistochemical methods. The expression of CK19 indicated that there were epidermal stem cell in cultured cells. 2, Organization culture methods were used , to acquired more pure fibroblasts. 3, CK19 was expressed in the outer root sheat of hair follicle, and in the stratum basale of glabrous skin. Conclusion: To successfully isolate, culture and subculture the keratinocytes and fibroblast. There are epidermal stem cell in the cultured keratinocyte. The epidermal stem cell localized in the outer root sheat of hair follicle,and in the stratum basale of glabrous skin.