Premium
Comparison of volumetric capillary cytometry with standard flow cytometry for routine enumeration of CD34+ cells
Author(s) -
Cabezudo E.,
Querol S.,
Cancelas J.A.,
Garcìa J.
Publication year - 1999
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1046/j.1537-2995.1999.39080864.x
Subject(s) - cd34 , flow cytometry , cord blood , leukapheresis , apheresis , enumeration , clonogenic assay , reproducibility , medicine , andrology , immunology , biology , chemistry , chromatography , platelet , stem cell , cell , mathematics , genetics , combinatorics
BACKGROUND: This study assesses the feasibility of a new volumetric cytometry system for the enumeration of CD34+ cells in apheresis components, peripheral blood, and cord blood samples in routine laboratory work. This system is compared with the following flow cytometry protocols: Milan, ISHAGE, ISHAGE with 7‐AAD, and flow‐count fluorospheres. STUDY DESIGN AND METHODS: Correlation, linearity, and reproducibility studies were performed for the various methods. Clonogenic cultures were performed, as an external control, to assess the correlation between the number of CD34+ cells per μL and the number of colony‐forming units per μL. RESULTS: The linear regression analysis demonstrated that the five methods were comparable (R 2 ranged from 0.86 to 0.96 and slopes were close to 1). The CD34+ assay and the flow‐count methods showed poor linearity for CD34+ cell counts below 10 cells per μL (R 2 = 0.46 and 0.47). The reproducibility assay for a CD34+ count of 10 cells per μL showed a CV of 12 percent and 25 percent for the Milan and CD34+ assay methods, respectively. The mean CV among all five methods for the 46 evaluated samples was 20 percent. There was a strong correlation between the number of CD34+ cells per μL and colony‐forming units per μL in cord blood and apheresis samples (r = 0.71‐0.81). CONCLUSION: The CD34+ assay is useful in CD34 enumeration in cord blood, leukapheresis samples, and peripheral blood samples and provides comparable results to the Milan, ISHAGE, ISHAGE with 7‐AAD, and flow‐count methods. Nevertheless, peripheral blood samples with low CD34 absolute counts (below 10 cells/μL) should be analyzed by alternative flow cytometry protocols. Even though the same operator performed the study in a single laboratory, the high inter‐method CV suggests that differences in sample preparation and gating strategy are factors that increase variability. Protocols with fewer intermediate steps or fully automated protocols such as the CD34+ assay are expected to reduced intra‐ and inter‐laboratory variability.