A reproducible procedure for primary culture and subsequent maintenance of multiple lines of human skin fibroblasts

Cultured fibroblasts are a valuable tool to study many cellular processes and their modification by aging. Fibroblasts are a useful cell type in which to study many diseases, including those of the nervous system, in which a strong genetic component is suspected. Fibroblasts permit the study of multiple, dynamic processes in living cells, while avoiding the effects of the dying process and post-mortem artifacts that limit other approaches. For results to be comparable across time in one laboratory or consistent between laboratories, the detailed culture techniques require meticulous care and replicability. Lack of attention to detail in initial stages can lead to selection of different cell populations. Small variations in other variables such as batches of serum can significantly alter growth rates and comparisons of cells from controls and Alzheimer patients. The aim of this paper is to present a detailed protocol for comparison of multiple cell lines from many patients. An example of using this approach to study growth and phase out (i.e., senescence) of cells from Alzheimer patients is presented. This procedure represents a modification of an earlier protocol (Cristofalo and Charpentier, 1980).