[Culture and identification of human skin fibroblasts].

ABSTRACT

OBJECTIVE: To explore the method of isolation, cultivation, and identification of human skin fibroblasts in vitro. METHODS: By digesting human skin with collagenase type II to isolate human eccrine sweat glands. The fibroblasts grew along with the growth of eccrine sweat gland cells,and they were separated by digesting with 0.25% trypsin and 0.02% ethylenediaminetetraacetic acid (EDTA). Dulbecco's modified Eagle's medium (DMEM) was the basic culture medium, being supplemented with fetal bovine serum (10%), penicillin (100 kU/L), and streptomycin(100 mg/L) Fibroblasts were incubated at 37 centigrade in a humidified atmosphere of 5% CO2 and 95% air in the incubator. Cellular morphologies were observed by inverted phase contrast microscopy and hematoxylin-eosin staining, and the cultured cells were identified by vimentin immunostaining and chromosome analysis. RESULTS: The isolated fibroblasts could grow and proliferate in vitro, and immunostaining of vimentin was positive in cultured fibroblasts and the number of chromosome was 46. CONCLUSION: Acquired human skin fibroblasts can be cultured in stable condition in vitro, and sufficient and reliable target cells can be obtained for the study of the mechanisms of wound healing at molecular level.