Approximately half of all CD45^midRho⁻EPCR⁺⁺ cells are HSCs, and these segregate into the same distribution of subtypes as CD45^midlin⁻Rho⁻SP HSCs. (A) The selection of a Rho⁻ population as defined by cells unstained with Rho. If the Rho⁻ cells are plotted in a CD45 vs EPCR plot (B), it is evident that the majority of the remaining EPCR⁺⁺ cells are CD45^mid and comprise a distinct population that represents only 0.11% of the Rho⁻ fraction for a total of 0.005% of the total adult mouse bone marrow cells. When 3 independent transplantation experiments were performed and the results were pooled, the frequency of HSCs was approximately 50% (C). Representative fluorescence-activated cell sorter plots for a repopulated mouse transplanted 16 weeks earlier with a single CD45^midRho⁻EPCR⁺⁺ cell are shown in panel D. By comparing the ratio of donor-derived B, T, and GM cells in the peripheral blood at 16 weeks, mice were determined to have been repopulated by one of the 4 previously defined HSC subtypes.¹⁰  The pie charts in panels E and F show the percentage of each subtype that is represented in each HSC population.