Figure 4.

Fractionation of human bone marrow (BM) using counterflow centrifugal elutriation combined with fluorescence-activated cell sorting using the five-antibody/viability marker panel shows that CD45⁻CD73⁺CD90⁺CD105⁺CD44⁻ MSPCs elute as small cells (70 and 90 ml/minute). (A–D): Representative display of SSC/FSC color density plots of the elutriated fractions 50/70 (A), 90 (B), 100–110 (C), and >110 (D). Display of CD45/7-AAD density plot was used to identify CD45⁻7-AAD⁻ (live) cells (gate R3), which was subgated onto a CD73/CD90 antigen plot and identified a cluster of live CD45⁻CD73⁺CD90⁺cells (gate R4), which was further subgated onto a CD105/CD44 antigen plot. Shown is the display of CD105⁺CD44⁻ on live CD45⁻CD73⁺CD90⁺ gated cells. The sort window shows a cluster of CD105⁺CD44⁻ cells (R5). The sorted live CD45⁻CD73⁺CD90⁺CD105⁺CD44⁻ cells (gate R5) are distributed close to the location of lymphocytes when back-gated onto an SSC-height/FSC-height dot plot (far right). Representative images from one experiment are shown. Similar results were seen in four other BM samples from different donors. Abbreviations: 7-AAD, 7-aminoactinomycin D; Frac, fraction; FSC, forward scatter; SSC, side scatter.