Expression of CD127 and Ig proteins on normal pediatric BM B-lineage cells and CD19⁺ cells from xenogeneic cultures. (A) CD127 expression was assayed on normal pediatric BM by flow cytometry. Total CD19⁺ cells in the lymphoid light-scatter gate were analyzed for surface μHC and CD34 expression (top right contour plot). CD127 expression was then analyzed and plotted on the 3 major populations of B-lineage cells in human BM: pro-B cells (CD19⁺/CD34⁺/μHC⁻), pre-B cells (CD19⁺/CD34⁻/μHC⁻), and immature/naive B cells (CD19⁺/CD34⁻/μHC⁺). CD127 expression was detected on all 3 subpopulations of B-lineage cells in human BM. (B) Cytocentrifuge slides of FACS-purified CD127⁺ and CD127⁻ cells from xenogeneic cultures were stained for expression of Igμ heavy chains using goat anti–human IgM TRITC (red) and counterstained with DAPI (blue). Mounting medium (2.3% wt/vol DABCO; Sigma D-2522/10% 1× PBS/87.7% glycerol) was used to cover the stained slides and slides were stored at 4°C and visualized at room temperature. Images were acquired using the Plan-Apochromat 10×/0.45 numerical aperture (NA) objective on an Axiovert 2 fluorescent microscope (Carl Zeiss) equipped with a Spot CCD camera (Diagnostic Instruments) and Spot Advanced 4.6 acquisition software. Adobe Photoshop 7.0 was used to create image overlays. Approximately 5% of cells in each population expressed Igμ. (C) CD19⁺ cells were isolated from 4-week xenogeneic cultures and stained for expression of CD127, Igμ, Igκ, and Igλ. (D) CD19⁺ cells were isolated from a 7-week xenogeneic cultures and stained for expression of Igμ and Igδ heavy chains.