Determination of the minimal number of BM-HSPCs, BM-MNCs and volume of BM-aspirate required for the reconstitution of NSG mice.

<p>Six-to eight week-old sublethally irradiated NSG mice were transplanted intra-femorally with non-cryoconserved CD34⁺ HSPCs (n = 14, 11 donors). (<b>A</b>) At >20 weeks after transplantation mice were analyzed by flow cytometry and engraftment in BM and spleen was determined by human CD45 expression and plotted vs. the number of injected HSPCs. The number of BM-HSPCs is calculated, which is required to achieve reconstitution in BM and spleen comparable to the lowest number of transplanted CB-HSPCs (4.8×10⁴ injected CD34⁺ HSPCs) that gives a reconstitution level matching with published data (BM: 6×10⁶ hCD45⁺; spleen 1.7×10⁶ hCD45⁺). Linear regression analysis revealed that at least 1.7–2.1×10⁵ BM-HSPCs are needed. (<b>B</b>) The number of BM-MNCs at sample receipt is plotted vs. the number of isolated HSPCs. The number of BM-MNCs (10⁸) is calculated that is required to isolate HSPCs sufficient for the reconstitution of six mice from one BM sample, given that each mouse receives 2.1×10⁵ HSPCs. (<b>C</b>) The number of CD34⁺ HSPCs per 10 million BM-MNCs is plotted vs. the age of carcinoma (n = 77) or control-patients (n = 7). No correlation is observed. (<b>D</b>) Percentage of samples and the volume of samples at receipt, which contained ≥10⁸ BM-MNCs allowing the isolation of HSPCs sufficient for the transplantation of six mice.