Cardiac stem cells were prepared with two methodologies. 1) Enzymatic dissociation of small myocardial samples (30mg), from which c-kit positive cells were sorted, using immunobeads and plated at low density to obtain multicellular clones from single founder cells. Lineage-negative (Lin-), c-kit-positive cells made up 41% of the population, and early committed cells (GATA4-positive) made up 59%.2 of the population. 2) Samples were cultured by the primary explant technique. Using this technique, 52% of the cells were Lin- and c-kit positive, and 48% were early committed cells.The culture medium for the second isolating methodolgy consists of F12 medium supplemented with FBS (5-10%) and insulin-selenium-transferrin mixture. In the presence of serum, hCSCs attach rapidly and continue to grow up to passage 8, undergoing 24 population doublings (PDs).