Characterization of the functional properties of human CB-derived HSC fractionated with CD166. (A) Human CB cells were analyzed with a combination of lineage markers, CD34 and CD38 and Lin⁻CD34⁺CD38⁻ cells (group 1) were analyzed for the expression of CD166 to identify CD34⁺CD38⁻CD166⁻ (group 2) and CD34⁺CD38⁻CD166⁺ (group 3) cells. Percentages next to each group represent the relative size of each population within the dot plot. (B) A total of 5000 cells from groups 1, 2, and 3 were transplanted into NSG mice and chimerism was assessed 4-months later (n = 3-4 per group). *P < .05, CD34⁺CD38⁻CD166⁺ vs other groups. (C) Fractionation of human CB cells with CD34 and a combination of lineage markers and CD38 to select Lin⁻CD34⁺CD38⁻ cells (group 1 in left dot plot). Lin⁻CD34⁺CD38⁻ cells were analyzed with CD166 and CD49f (right dot plot) to define groups 2, 3, 4, and 5. Percentages next to each group represent the relative size of each population within the dot plot. A total of 1000 cells from groups 1 through 5 were transplanted into NSG mice. (D) BM chimerism 4-months PT |(n = 3-4 per group). *P < .05, groups 2 and 3 vs other groups. Similar results were obtained in another independent experiment (4000 cells/group; n = 4/group). The 2 data sets were not pooled. (E-F) Chimerism at 4-months PT (E) sustained by 1000 cells per mouse from groups 1 to 4, as shown in panel (C) and in secondary recipients (F) transplanted with BM cells collected from primary recipients (one-half a femur equivalent from pooled BM cells per recipient) shown in (E) (n = 4-5 per group in [E] and [F]). *P < .05, groups 2 and 3 vs other groups. (F) Flow cytometric analysis of representative mice from (F) are shown in supplemental Figure 10.