Isolation, culture and, characterization of Cardiosphere-Derived Cells (CDCs)

Abstract Cardiosphere-derived Cells (CDCs) can be isolated from cardiac tissues of pigs. The protocol begins with an enzymatic digestion followed by a pre-culture in a specific cell culture medium. Adherent cells from digested tissue are detached and seeded with the Cardiosphere Growing Medium (CGM). Under these conditions, cells clusters are formed and migrating cells are selected and seeded again to expand CDCs. Phenotypical, functional and molecular assays are necessary for the characterization of isolated cells. Phenotypic analysis is performed by flow cytometry to identify surface markers. For functional analysis, cells are in vitro differentiated using different culture media, then stained and observed by optical microscopy. Finally, PCR can be used to analyse the gene expression profile.This protocol is suitable for establishing a CDCs culture for multiples applications: in vitro assays, preclinical studies, “omics” analyses, etc.