Efficient isolation and enrichment of mesenchymal stem cells from bone marrow

Abstract

Bone marrow (BM) mesenchymal stromal cells (MSC) have been identified as a source of pluripotent stem cells used in clinical practice to regenerate damaged tissues. BM MSC are commonly isolated from BM by density-gradient centrifugation. This process is an open system that increases the risk of sample contamination. It is also time consuming and requires technical expertise that may result in variability regarding cellular recovery. The BD Vacutainer® Cell Preparation Tube™ (CPT) was conceived to separate mononuclear cells from peripheral blood. The main goal of this study was to verify whether MSC could be isolated from BM using the CPT. BM was harvested, divided into two equal aliquots and processed using either CPT or a Ficoll-Paque™ PREMIUM density gradient. Both methods were compared regarding cell recovery, viability, proliferation, differentiation capacities and the presence of MSC progenitors. Similar numbers of mononuclear cells were isolated from BM when comparing the two methods under study. No differences were found in terms of phenotypic characterization, viability, kinetics and lineage differentiation potential of MSC derived by CPT or Ficoll. Surprisingly, a fibroblast-colony-forming unit (CFU-F) assay indicated that, with CPT, the number of MSC progenitors was 1.8 times higher compared with the Ficoll gradient separation. The CPT method is able to isolate MSC efficiently from BM, allowing the enrichment of MSC precursors.