Parameters that influence the isolation of multipotent mesenchymal stromal cells from human umbilical cord blood

Background and objectives Umbilical cord blood is an important source of stem cells. However, isolating multipotent mesenchymal stromal cells (MSCs) from umbilical cord blood presents methodological challenges. We compared the effectiveness of six approaches to improve the success rate of MSC isolation and proliferation from umbilical cord blood. Methods Thirty umbilical cord blood units underwent investigation. In 10 samples, MNCs from each sample were divided into four groups to test the effect of negative immunodepletion (NI) alone (group A); NI plus basic fibroblastic growth factor (bFGF) supplementation together (group B); bFGF supplementation alone (group C); and culture with neither NI nor bFGF (group D). The cells of each group were isolated from 10 mL of umbilical cord blood. For investigating the effect of sample volume (group E) and MesenCult Proliferation Kits (group F), cells were isolated from 45 ± 2 ml. MSCs were identified on the basis of morphological, flow cytometric and differentiation potential characteristics. Results In groups of A–D, one week after the initial seeding, the cells showed a rounded appearance, and in the fourth week, many of them died. MSCs outgrowth was seen in 40% of the samples from group F, and this yield was further enhanced to 60% in cultures done with the MesenCult Proliferation Kit (group F). The fibroblast-like cells expanded rapidly and showed features of MSCs. Conclusion Sample volume was the parameter that showed the greatest influence on the isolation yield of MSCs from umbilical cord blood. This could be further enhanced by adding the MesenCult Proliferation Kit.