An Optimized Method to Isolate Human Fibroblasts from Tissue for Ex Vivo Analysis

Despite their involvement in many physiological and pathological processes, fibroblasts remain a poorly-characterized cell type.Analysis of primary fibroblasts while maintaining their in vivo phenotype is challenging: standard methods for fibroblast isolation require cell culture in vitro, which is known to alter phenotypes.Previously-described protocols for the dissociation of primary tissues fail to extract sufficient numbers of fibroblasts, instead largely yielding immune cells.Here, we describe an optimized method for generating a fibroblast-enriched single-cell suspension from human tissues using combined mechanical and enzymatic dissociation.This allows analysis of ex vivo fibroblasts without the need for culture in vitro.