Cell sorting of transplanted cell populations. (A) Cells were obtained for immunoglobulin sequencing. At 8 weeks after transplantation, cells were collected from the spleen and bone marrow of chimeric mice that had received transplants of CD34⁺ lymphoid progenitors (in the case of fetal and adult bone marrow) or cord blood mononuclear cells. Cells were enriched for human cells by depleting mouse myeloid and macrophage lineage positive cells. CD34^-CD19⁺ cells (left box) were sorted into surface IgM^- CD5^- (i), IgM⁺ CD5^- (ii), or IgM⁺ CD5⁺ (iii) populations (right box) in the bone marrow and spleen (the latter with the exception of surface IgM^- cells). (B) Anti-V gene antibodies were used to determine the contribution of the VH4 family to the chimeric repertoire. Surface IgM⁺ cells (left) were stained with monoclonal antibodies that together recognize all VH4 family members. 9G4 recognizes VH4-34 immunoglobulins and LC1 recognizes all VH4 family members except VH4-34. Thus 17.6% of the repertoire was LC1 positive and 5.5% of the repertoire was positive for 9G4 binding, which combined equals 23.1% of cells that express an immunoglobulin using a member of the VH4 family. These monoclonal antibodies were also used to determine the contributions of VH4-34 (recognized by 9G4) relative to the other VH4 family members (recognized by LC1). 9G4 staining cells comprised 23% of cells that stain for any member of the VH4 family (not related to the percentage of VH4 staining cells among the entire cell population). (C) CD34^- cells from adult bone marrow do not stably engraft NOD/SCID mice. Mice received transplants of either adult bone marrow CD34⁺ cells (left) or cord blood CD34^- cells (right). Engraftment was compared by screening the peripheral blood first for human CD45⁺ cells followed by CD19 and CD3.