Harold W. Fisher

The parental HeLa cell population, a morphologically uniform, human cancer cell strain, grown for several years in tissue culture by procedures always involving massive inocula, has been shown to contain different mutant cell types. Two clonal lines have been isolated and established as reliable stock cultures. Both strains exhibit 100 per cent plating efficiency in high or low serum concentrations in the presence of a feeder system. In the absence of a feeder system and in low serum concentrations, the two strains are quantitatively differentiable: S3 still exhibits 100 per cent plating efficiency, while that of S1 lies in the neighborhood of zero. These differences have remained stable throughout 100 successive generations of growth of each strain including 2 single cell isolations. Application of these techniques to studies in the genetics of mammalian somatic cells and to specific cell-cell interactions has been indicated.

A methodology has been described for reliable cultivation in vitro of dispersed fibroblastic cells obtained from normal human organs. The procedure has permitted establishment of stable cell lines from almost every sample taken, among which the following organs were represented: skin, spleen, amnion, lung, liver, bone marrow, brain, muscle, and heart. Equally good growth has been achieved with cells from embryonic or adult tissues. The methods previously developed whereby single cells plated in Petri dishes grow into isolated macroscopic colonies can successfully be applied to the plating of human fibroblastic stocks. Plating efficiencies in the neighborhood of 50 to 60 per cent are readily achieved with such strains. The resulting colonies can be picked and clonal stocks established. Fibroblastic morphology is maintained in the colonies arising from every single cell of such clonal stocks. All of the single cells from epithelioid clonal strains also maintain their integrity throughout repeated subculture. Since the difference between clonal stocks of these two types is always maintained whenever the respective single cells are plated in the same medium, regardless of the previous history of these stocks, it may be concluded that a true genetic difference exists in these cell lines. In addition to the morphological differences between epithelioid and fibroblastic cell strains, the latter have more demanding nutritional requirements for single cell growth. Thus, single cells of fibroblastic lines almost never produce colonies with high efficiency unless the growth medium which is sufficient for epithelioid cells is supplemented with embryo extract, or a cell feeder layer. Fibroblastic cells are also more resistant to tryptic digestion of the bond uniting the cells to glass surfaces. By use of differential media, growth of both fibroblastic and epithelioid cells, respectively, has been obtained, from dispersed single cells obtained by trypsinization of a specimen of human embryonic lung.

Proceedings of the National Academy of Sciences (PNAS), a peer reviewed journal of the National Academy of Sciences (NAS) - an authoritative source of high-impact, original research that broadly spans the biological, physical, and social sciences.

Contact inhibition of replication of the established mammalian cell line 3T3 was examined during growth of the colony and compared with that of the Chinese-hamster cell line CHL-1. The growth curves of cells in the colonies conformed to the predicted exponential and linear rates for CHL-1 and 3T3 respectively. Autoradiographs of colonies in which DNA was labeled with tritiated thymidine showed that in 3T3 colonies, only peripheral cells were labeled, while CHL-1 colonies were labeled throughout.