Abstract
Quantitating CD34 antigen-positive (CD34+) cells by flow cytometry is a preferred method for assessing the graft adequacy of peripheral blood stem cell (PBSC) collections. It has been reported that the CD34+ population, however, differs among several counting methods depending on the protocol used. Here, we analyzed a total of 131 samples of peripheral blood (n=75), PBSC (n=40) and frozen-thawed PBSC (n=16) for %CD34+ cells and the absolute number of CD34+ cells using single-platform and two-platform protocols with the Cytomics FC500 and Facs Calibur.
%CD34+ cells showed high correlation coefficients (R2=0.980 for peripheral blood, 0.979 for PBSC, and 0.796 for frozen-thawed PBSC) between the Cytomics FC500 and Facs Calibur, and the absolute number of CD34+ cells did not differ between the two machines. For PBSC and frozen-thawed PBSC samples, the absolute number of CD34+ cells correlated well between the single-platform and two-platform methods using the Cytomics FC500 and Facs Calibur. The number of CD34+ cells in peripheral blood detected by the two flow cytometers was, however, higher (Cytomics FC500 : p=0.005, Facs Calibur : p=0.0004) by the two-platform than the single-platform method.
These results show that results for CD34+ cell numbers using the Cytomics FC500 are similar to those with the Facs Calibur when the single-platform method is used. Further, the variation in CD34+ cell count among operators was dependant on errors attributed to pipetting at the dilution of samples or at the addition of beads for CD34+ cell assay. The degree of error attributed to pipetting during dilution with the two-platform and single-platform methods was not significantly different.
Variation in CD34+ cell counts can be reduced by using a single-platform technique in accordance with the International Society of Haematotherapy and Graft Engineering guidelines.
