ISOLATION AND CHARACTERIZATION OF CD34+ HEMATOPOIETIC STEM-CELLS FROM HUMAN PERIPHERAL-BLOOD BY HIGH-GRADIENT MAGNETIC CELL SORTING

CD34+ cells, present at a frequency of 0.18 +/- 0.052% among leukocytes from peripheral blood (PB), can be rapidly and efficiently (recovery of 39.0-74.0%) enriched to a frequency of 38.6-87.1% (54.4 +/- 12.3%) by high-gradient magnetic cell separation (MACS) for immunophenotyping, characterization in colony-forming cell (CFU) assays, and further purification to homogeneity (>98%) by multiparameter fluorescence-activated cell sorting (FA(CS). Enriching PB-CD34+ cells for immunophenotyping allows the detection of small subpopulations, expressing the B-cell antigens CD10, CD19, and CD20, the T-cell antigens CD45RA and CD7, and a small subpopulation expressing high levels of CD34 (1.20 +/- 0.12%), which mostly coexpress CD19 (91.9 +/- 9.05%), CD20 (64.8 +/- 14.4%), and CD38 (84.5 +/- 10.3%). All PB-CD34+ cells express elevated levels of CD71 (transferrin receptor), with a subpopulation of high expressing cells, and CD38. Some cells express CD33. MACS-enriched PBCD34+ cells show ''normal'' hematopoietic colony formation in vitro. The ease and efficiency of purification of large numbers of CD34+ cells from PB by MACS is not only relevant for the characterization of migrating stem cells but also opens new possibilities for stem cell transplantation and genetic manipulation of the hematopoietic system.