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Isolation of CD34+ progenitor cells from peripheral blood by use of an automated immunomagnetic selection system: factors affecting the results
Authors:Martín-Henao G A  Picón M  Amill B  Querol S  González J R  Martínez C  Martino R  Ferrá C  Brunet S  Grañena A  Sierra J  García J
Institution:Department of Cryobiology and Cell Therapy, Cancer Research Institute, Barcelona, Spain. gmartinh@iro.es
Abstract:BACKGROUND: The isolation of CD34+ cells from mobilized peripheral blood is being increasingly used in the setting of allogeneic or autologous hematopoietic cell transplantation. Investigation of variables that may influence the effectiveness of CD34+ cell selection is of interest. STUDY DESIGN AND METHODS: Fifty-one CD34+ cell selections from peripheral blood progenitor cells (PBPCs) (39 allogeneic and 12 autologous) were performed using a magnetic cell separator (Isolex 300i, Baxter), including version 2.0 software. The results obtained were analyzed for different processing variables. The feasibility of transplanting these isolated CD34+ cells was also analyzed. RESULTS: The isolated CD34+ cell fraction had a median purity of 88.9 percent (range, 47.8-98.3). The median recovery of CD34+ cells was 45.1 percent (13.8-76.2), and the median colony-forming unit- granulocyte-macrophage (CFU-GM) content was 17. 2 percent (0.8-58.6). Logarithms of T- and B-cell depletion had median values of 3.7 and 2.8, respectively. The version 2.0 software of the Isolex 300i gave a higher CD34+ cell recovery in the enriched cell fraction (median 57.8%) than did version 1.11 (39.4%) or 1.12 (44.4%) (p = 0.01). The use of recombinant human deoxyribonuclease I during cell processing yielded more CD34+ cells (53% vs. 41%, p = 0. 01) and higher purity (92.8% vs. 87%, p = 0.03). There was a correlation between the percentage of CD34+ cells labeled with the monoclonal antibody 8G12 clone and the percentage of CD34+ cells labeled with the monoclonal antibody used during the processing technique (9C5 clone) in the initial, enriched, and depleted CD34+ cell fractions (R(2) = 0.95; 0.92; 0.78, p< 0.005, respectively). Median times for recovering >0.5 x 10(9) per L of granulocytes and >20 x 10(9) per L of platelets were 13 and 16 days in the allograft patients and 13 and 14 days in the autograft patients. CONCLUSION: CD34+ cells can be highly and effectively isolated from allogeneic and autologous grafts by use of this automated technique, with a high grade of T- and B-cell depletion. These purified CD34+ cell components can engraft normally.
Keywords:7-AAD = 7-aminoactinomycin-D  CFU = colony-forming units  CSF = colony-stimulating factor  DNase = deoxyribonuclease I  FITC = fluorescein isothiocyanate  GAM = goat anti-mouse  GM = granulocyte  IB(s) = immunomagnetic bead(s)  LK = leukapheresis  MFI = mean fluorescence intensity  MoAb(s) = monoclonal antibody(ies)  NC(s) = nucleated cell(s)  PBPC(s) = peripheral blood progenitor cell(s)  PE = phycoerythrin  PR = peptide release reagent  rHu = recombinant human
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