Transplant centers often rely on CD34+ cell quantitation by flow cytometry to ensure adequacy of hematopoietic progenitor cell collection. Because of variation in interpretation, a lack of interlaboratory proficiency studies, and no generally accepted methodology, comparison of CD34 data from site to site is difficult. Twenty-one samples from marrow and peripheral blood stem cell collections were shipped to 10 participating North American laboratories for analysis. Duplicate samples were included to assess reproducibility. Participants were surveyed for methodology. Three centers had previously attempted to standardize their methodology among themselves. The variability observed in the CD34 values ranged from a max/min. reported value per sample of 2.9 to 749 (median 76). Exclusion of two outlying sites reduced the variability of results to 1.2 to 27 (median 3.1). Variation among the three standardized sites ranged from 1.2 to 4.4 (median 1.6). Overall reproducibility (excluding the outlying sites B and G) ranged from a minimum of 0-16.5 (percent mean difference) for site C to a maximum of 4.1-133 for site H. Strategies for gating were found to largely influence results. We observed an alarming variation among the CD34 cell counts reported from different laboratories. Standardization substantially reduced observed variation. The need for standardized methodology, reporting, quality control, and proficiency testing is underscored by these findings.
Accurate enumeration of CD34+ stem cells is important in assessing the need for continued mobilization and subsequent apheresis collections. We compared two new analysis systems, ProCOUNT (Becton Dickinson Immunocytometry Systems) and IMAGN 2000 STELLer (Biometric Imaging, Inc.) with our current (3-Color) flow cytometry-based method. The ProCOUNT system uses an absolute counting tube, which contains reference beads and a specific (multiple) gating strategy to determine an absolute count. The STELLer assay combines microvolume fluorimetry and automated analysis software to determine an absolute count. To evaluate linearity and reproducibility, peripheral blood was spiked with CD34+ cells (KG1a cell line). Three dilution series (measured at approximately equal to 0, 5, 10, 25, 50, and 100 CD34+ cells/microliter) were analyzed by each method. Analysis of predicted versus actual CD34+ concentration showed excellent correlation with all methods (r2 > or = 0.97, slope 0.98-1.04). To further assess precision, two PBSC samples, at approximately 200 and 800 CD34+ cells/microliter, respectively, were analyzed 10 times by each method. Coefficients of variation for the precision analysis of these samples were 5.1%-6.4% and 5.4%-12.3%, respectively. To assess overall performance, 75 patient specimens were analyzed. Excellent correlation (r2 values of 0.89-0.98) was observed among all three methods. We conclude that the three methods provide comparable linearity and reproducibility.
Benjamin et al.' recentlyreported inTRANSFUSION a poor correlation between circulating CD34+ cell counts and subsequent CD34+ peripheral blood progenitor cell (PBPC) yields. Such an observation is contrary to several other recent reports and our own experience (Fig. 1) that indicate a strong relationship between the number of circulating CD34+ cells and the CD34+ cell yield in apheresis-derived PBPC component^.^*^ Such comparisons are superior to those obtained with other surrogate measures such as circulating mononuclear cell count (Fig. 2). CD34+ cells collected (xl 06/kg)Fig. 1. Plot of circulating CD34+ cells versus CD34+ PBPC yield (x106) per kg in 38 collections (19 patients, 2 collections each). An excellent correlation was observed (P = 0.95). A virtually identical relationship was also observed in the plot of circulating CD34+ versus CD34+ PBPCs collected (x106/kg/L of blood processed) (data not shown). 81 / I k I . m I / L d 0 1 b ~ Q) Q a 0 10 20 30 40 J= 01 .b CD34+ cells collected (xl 06/kg) Fig. 2. Plot of circulating mononuclear cell count versus CD34+ PBPCs yield x 106/kg in 38 collections (19 patients, 2 collections each). A poor correlation was observed (I = 0.28). A virtually identical relationship was also observed In the plot of circulating mononuclear cell count versus CD34+ PBPCs collected (x106/kg/L of blood processed) (data not shown).Although it is unclear why Benjamin et al. found such a poor correlation, difficulties with enumeration of low numbers of CD34+ cells are well d o~u m e n t e d .~One possible explanation for the poor correlation may be imprecision (lack of reproducibility) in a method that fails to count at least 20,000 evenk4The number of events that Benjamin et al. counted was not indicated.The ability to accurately predict the optimal time to initiate PBPC harvest is important to maximizing the use of expensive medical resources (PBPC collection and cryopreservation) and minimizing the amount of cryoprotectant that would ultimately be returned to the patient. New, rapid, and relatively simple enumeration technologies are now becoming a~ailable.~ Such technologies will facilitate the accurate timing of PBPC collections. REFERENCESBenjamin RJ, Linsley L, Fountain D, et al. Preapheresis peripheral blood CD34+ mononuclear cell counts as predictors of progenitor cell yield. Transfusion 1997;37:79-85. Mohle R, Murea S, Pforsich M, et al. Estimation of the progenitor cell yield in a leukapheresis product by previous measurement of CD34+ cells in peripheral blood. Vox Sang Elliot BC, Samson DM, Armitage S, et al. When to harvest peripheral blood stem cells after mobilization therapy: prediction of CD34-positive cell yield by preceding day CD34positive concentration in peripheral blood. J Clin Oncol Brecher ME, Sims L, Schmitz J, et al. North American multicenter study on flow cytometric enumeration of CD34+ hematopoietic stem cells. J Hematother 1996:5;227-36. Sims L, Brecher ME, Gertis K, et al. Enumeration of CD34positive stem cells: evaluation and comparison of methods.The ab...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.