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Evaluation of a New Single-Parameter Volumetric Flow Cytometer (CyFlow^green) for Enumeration of Absolute CD4⁺ T Lymphocytes in Human Immunodeficiency Virus Type 1-Infected Thai Patients

Center of Excellence for Flow Cytometry, Office for Research and Development, Faculty of Medicine, Siriraj Hospital, Mahidol University, Bangkok, Thailand,¹ Thailand Ministry of Public Health-U.S. CDC Collaboration, Nonthaburi, Thailand,² Bureau of AIDS, TB and STIs, Department of Diseases Control, Ministry of Public Health, Nonthaburi, Thailand,³ Division of HIV/AIDS Prevention, Centers for Disease Control and Prevention, Atlanta, Georgia,⁴ Global AIDS Program, Centers for Disease Control and Prevention, Atlanta, Georgia⁵

Received 14 May 2005/ Returned for modification 29 July 2005/ Accepted 16 September 2005

Use of the standard dual-platform flow cytometric method for determination of CD4⁺ T-lymphocyte counts, which needs both a flow cytometer (FCM) and hematological analyzer, would inevitably lead to increased variability. The development of new single-platform (SP) FCMs that provide direct CD4⁺ T-lymphocyte counts for improved assay precision and accuracy have recently attracted attention. This study evaluated one of those systems, CyFlow^green (Partec), a single-parameter SP volumetric FCM. The performance of CyFlow^green was compared with those of two reference standard SP microbead-based technologies of the three-color TruCOUNT tube with the FACScan FCM and a two-color FACSCount system (Becton Dickinson Biosciences). Absolute CD4⁺ and CD8⁺ T-lymphocyte counts in 200 human immunodeficiency virus type 1-seropositive blood specimens were determined. Statistical analysis for correlation and agreement were performed. A high correlation of absolute CD4 counts was shown when those obtained with CyFlow^green were compared with those obtained with the bead-based three-color TruCOUNT system (R² = 0.96; mean bias, –69.1 cells/µl; 95% confidence interval [CI], –225.7 to +87.5 cells/µl) and the FACSCount system (R² = 0.97; mean bias, –40.0 cells/µl; 95% CI, –165.1 to +85.1 cells/µl). The correlation of the CD4⁺ T-lymphocyte counts obtained by the two bead-based systems was high (R² = 0.98). Interestingly, CyFlow^green yielded CD4⁺ T-lymphocyte counts that were 21.8 and 7.2 cells/µl lower than those obtained with the TruCOUNT and the FACSCount systems, respectively, when CD4⁺ T-lymphocyte counts were <250 CD4⁺ T-lymphocyte counts/µl range or 17.3 and 5.8 cells/µl less, respectively, when CD4⁺ T-lymphocyte counts were <200 cells/µl. The single-parameter CyFlow^green volumetric technology performed well in comparison with the performance of the standard SP bead-based FCM system. However, a multicenter comparative study is needed before this FCM machine is implemented in resource-limited settings.