Direct volumetric flow cytometric quantitation of CD34+ stem and progenitor cells

• Published in: Transfusion medicine (Oxford, England) England), 2012-06, Vol.22 (3), p.205-210
• Main Authors: Gutensohn, K., Nikolitsis, A., Gramatzki, M., Spitzer, D., Buwitt-Beckmann, U., Humpe, A.
• Format: Article
• Language: English
• Subjects: Adult; Aged; Antigens, CD34 - analysis; bead technology; Blood Cell Count - methods; Blood Component Removal; Blood Preservation; CD34; Cryopreservation; Female; flow cytometry; Flow Cytometry - instrumentation; Flow Cytometry - methods; Hematopoietic Stem Cell Mobilization; Hematopoietic Stem Cell Transplantation - methods; Hematopoietic Stem Cells - chemistry; Humans; Leukocyte Common Antigens - analysis; Male; Microspheres; Middle Aged; Peripheral Blood Stem Cell Transplantation - methods; Reproducibility of Results; single platform; volumetric analysis
• ISSN: 0958-7578; 1365-3148
• DOI: 10.1111/j.1365-3148.2012.01155.x

Objectives: In this study, we compared a classic single‐platform (SP) method applying beads for enumeration of CD45+ or CD34+ cells with a new device allowing direct volumetric measurements of stem and progenitor cells. Background: Following apheresis and cyropreservation, the precise enumeration of CD34+ cells as key parameter of graft quality is mandatory for the clinical course after transplantation. Currently, flow cytometry with SP technique represents the ‘gold standard’ for such determinations. Methods/Materials: Fresh samples, 14 from mobilised peripheral blood (PB), 9 from apheresis products (AP) and 13 samples from frozen‐thawed (FT) haematopoietic progenitor cell grafts, were analysed for CD34+ cells, CD45+ cells, and in frozen‐thawed samples for viability by a bead‐based flow cytometric method and in parallel by a direct, volumetric flow cytometric method. Results: Comparison of CD34+ analyses revealed a significant correlation (P < 0·01) for each material between both techniques with r = 0·95 (PB), r = 0·933 (AP) and r = 0·929 (FT). Also, for analysis of CD45+ cells µL−1, the measured numbers evaluated with the different techniques did not significantly differ for all three materials analysed. In frozen‐thawed samples, the analysis of viability was comparable for both techniques. Conclusions: The results of this study demonstrate that a direct volumetric analysis of CD34+ cells µL−1 or CD45+ cells µL−1 is feasible. This technique represents a simple and economical approach for standardisation of progenitor and stem cell analyses.