title: Separation of myeloid and erythroid progenitors based on expression of CD34 and c-kit.
abstract: In this report, a novel approach is described to physically separate erythroid progenitors from monocyte and granulocyte progenitors, based on the expression of CD34 and Kit. Using biotin-labeled human Kit ligand (KL) and flow cytometry, Kit was detectable on 2% to 3% of the nucleated cells in rhesus monkey bone marrow. Combination of biotin-KL with CD34 monoclonal antibodies (MoAb) showed that Kit was expressed on subsets of CD34low and CD34pos cells. Our data clearly demonstrate that CD34pos cells are more heterogeneous with respect to Kit expression than observed in studies using Kit MoAb. A small cluster, approximately 7% of the CD34pos cells, expressed CD34 at submaximal levels and stained brightly with biotinylated KL. This CD34pos/kithi fraction contained predominantly erythroid progenitors (burst-forming units-erythroid; BFU-E). The majority of the granulocytic and monocytic progenitors (colony-forming units-granulocyte/macrophage; CFU-GM) were CD34pos/kitmed. Some BFU-E were also detected in the CD34pos/kitmed and CD34low/kitpos fractions at low frequency. In the latter subset, most erythroid colony-forming units (CFU-E) were recovered. Using three-color flow cytometry, we analyzed expression of Kit in relation to that of CD34 and the class II major histocompatibility antigen, RhLA-DR. The most immature bone marrow cells that can be identified in vitro, ie, CD34pos/RhLA-DRlow cells, were kitmed. The CD34pos/kithi and CD34pos/kitneg subsets predominantly contained the more mature RhLA-DRbright cells. Our results demonstrate that erythroid precursors express c-kit at much higher levels than monomyeloid precursors and pluripotent progenitors. The difference in expression levels of CD34 and c-kit can be exploited to isolate BFU-E populations that are virtually devoid of nonerythroid cells.
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002469:Cell Separation / Q000379:methods*; D003114:Colony-Forming Units Assay; D015672:Erythroid Precursor Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D005434:Flow Cytometry; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006098:Granulocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D006410:Hematopoiesis / Q000187:drug effects; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology; D008253:Macaca mulatta; D009000:Monocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism*; D019089:Stem Cell Factor / Q000378:metabolism
chemical_list: D018952:Antigens, CD34; D007377:Interleukin-3; D019089:Stem Cell Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D019009:Proto-Oncogene Proteins c-kit