Identification and comparison of CD34-positive cells and their subpopulations from normal peripheral blood and bone marrow using multicolor flow cytometry

Abstract

Four-color flow cytometry was used with a cocktail of antibodies to identify and isolate CD34+ hematopoietic progenitors from normal human peripheral blood (PB) and bone marrow (BM). Mature cells that did not contain colony forming cells were resolved from immature cells using antibodies for T lymphocytes (CD3), B lymphocytes (CD20), monocytes (CD14), and granulocytes (CD11b). Immature cells were subdivided based on the expression of antigens found on hematopoietic progenitors (CD34, HLA-DR, CD33, CD19, CD45, CD71, CD10, and CD7). CD34+ cells were present in the circulation in about one-tenth the concentration of BM (0.2% v 1.8%) and had a different spectrum of antigen expression. A higher proportion of PB-CD34+ cells expressed the CD33 myeloid antigen (84% v 43%) and expressed higher levels of the pan leukocyte antigen CD45 than BM-CD34+ cells. Only a small fraction of PB-CD34+ cells expressed CD71 (transferrin receptors) (17%) while 94% of BM-CD34+ expressed CD71+. The proportion of PB-CD34+ cells expressing the B-cell antigens CD19 (10%) and CD10 (3%) was not significantly different from BM-CD34+ cells (14% and 17%, respectively). Few CD34+ cells in BM (2.7%) or PB (7%) expressed the T-cell antigen CD7. CD34+ cells were found to be predominantly HLA-DR+, with a wide range of intensity. These studies show that CD34+ cells and their subsets can be identified in normal PB and that the relative frequency of these cells and their subpopulations differs in PB versus BM.