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Background aimsThe manufacture of multipotential stromal cell (MSC)-based products is costly; therefore, a rapid evaluation of bone marrow (BM) ‘quality’ with respect to MSC content is desirable. The aim of this study was to develop a rapid single-platform assay to quantify MSC in BM aspirates.MethodsAspirated MSC were enumerated using the CD45−/low CD271bright phenotype and AccuCheck counting beads and compared with a classic colony-forming unit–fibroblast (CFU-F) assay. The phenotype of CD45−/low CD271bright cells was defined using a range of MSC (CD73, CD105, CD90) and non-MSC (CD31, CD33, CD34, CD19) markers. The effect of aspirated BM volume on MSC yield was also determined.ResultsCD45−/low CD271bright cells had a classic MSC phenotype (CD73+ CD105+ CD90+ ). Their numbers correlated positively with CFU-F counted manually (R=0.81, P

Aspiration of iliac crest bone marrow (ICBM) remains the most frequent technique used in harvesting multipotential stromal cells (MSCs) for bone regeneration. Although this tissue type is easily accessed by a surgeon, it has a low frequency of MSCs, which is significant given the high cell numbers required for bone regeneration strategies. Lipoaspirates possess higher MSC frequencies, albeit cells with a differentiation profile less suited to orthopaedic interventions. Intra-medullary cavities of long bones have previously been shown to harbour MSCs in animals, however evaluation of their frequency, differentiation capacity and phenotype in humans had not previously been performed. Long bone fatty bone marrow (LBFBM) was collected prior to harvesting bone graft. Basic cellular compositions of donor-matched LBFBM and ICBM aspirates, including the numbers of CD34+ hematopoietic stem cells and CD31+ endothelial cells, were similar. MSCs were enumerated using colony-forming-unit-fibroblast assays and flow cytometry for the presence of a resident LBFBM CD45−/low CD271+ MSC population and revealed a trend for higher MSC numbers (average 5 fold, n = 6) per millilitre of LBFBM compared to donor-matched ICBM. Functional characteristics of resident MSCs, including their growth rates, differentiation potentials and surface phenotypes (CD73+CD105+CD90+) before and after culture-amplification, were similar. Enhanced numbers of MSCs could be recovered following brief enzymatic treatment of solid fragments of LBFBM. Our findings therefore reveal that the intramedullary cavity of the human femur is a depot of MSCs, which, although closely associated with fat, have a differentiation profile equivalent to ICBM. This anatomical site is frequently accessed by the orthopaedic/trauma surgeon and aspiration of the intramedullary cavity represents a ‘low-tech’ method of harvesting potentially large numbers of MSCs for regenerative therapies and research. This article is part of a Special Issue entitled: Interactions Between Bone, Adipose Tissue and Metabolism.