Establishment, Maintenance, and Cloning of Human Dermal Fibroblasts

The widespread use of human diploid fibroblasts in many tissue-culture-based systems has its origins in the pioneering work on cellular senescence by Hayflick (). He established a reliable protocol for the maintenance of fibroblast strains that was also favorable for stimulating cell proliferation. The reproducibility of the cell cultivation system he developed allowed the realization that normal diploid cells exhibit a limited proliferative potential in vitro. Depending on the age of the donor and the biopsy site, human dermal fibroblasts can reach over 50 population doublings before senescence sets in (). This high value is of great advantage to laboratories wishing to study some aspect of cell function in a normal diploid cell population. With the development of reliable cryopreservation protocols, it is possible to build up a substantial bank of early to mid passage fibroblasts from a single primary culture derived from a biopsy. This bank of stable diploid cells thus has many attributes usually only found in transformed cell lines where homogeneity of cell population over a prolonged period of time is often crucial to the experimenter. Unlike a transformed line however, human fibroblasts maintained, as described here, generally retain the normal diploid karyotype.