Coetzee L.M., Tay S.S., Lawrie D., Janossy G., Glencross D.K.
Department of Molecular Medicine and Haematology, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; Collaborative Transplantation Research Group, University of Sydney, Sydney, NSW, Australia
Coetzee, L.M., Department of Molecular Medicine and Haematology, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; Tay, S.S., Collaborative Transplantation Research Group, University of Sydney, Sydney, NSW, Australia; Lawrie, D., Department of Molecular Medicine and Haematology, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; Janossy, G., Department of Molecular Medicine and Haematology, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa; Glencross, D.K., Department of Molecular Medicine and Haematology, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa
Background: CD38 expression on CD8+ T lymphocytes in HIV-infected patients is monitored by flow cytometry (FCM). There is however no consensus re CD38 protocols, analyses or result reporting within/ between laboratories. Internal quality control measures (QC) were established for a standardized CD38 protocol and a system proposed for reporting CD38 fluctuation in longitudinal HIV+ patient monitoring. Methods: A single-platform (SP) CD38/CD8 protocol was "piggy-backed" onto the standardized "panleucogating" CD45/CD4+ protocol. A weekly QC was established to monitor instrument stability (Flow-SETTM) and absolute cell count accuracy and reproducibility (stabilized blood product, Immuno-TrolTM). The Mean Fluorescence Intensity (MFI) of CD38 expression on CD8+-lymphocytes was monitored on both stabilized blood and HIV-control samples. Linearized MFI values were determined from biological controls, i.e. healthy donor monocytes and granulocytes, and tested as a method of reporting CD38 expression on selected HIV+ patients on ART. Results: The CD45/CD4/CD8/CD3 method for lymphocyte enumeration compared well with the CD38 protocol (CD45/CD4/CD8/CD38) with excellent similarity (±100%) and precision for absolute CD4 and CD8 counts (CVs < 5%). Fluorosphere MFI- (FlowSetTM, FlowCountTM) and color compensation values were exceptionally stable over time. CD38 MFI values established on monocytes as biological control was 4.0 and <2.0 for HIV-control lymphocytes. Conclusions: Monitoring FCM with fluorosphere MFI values, color compensation, and biological controls, can ensure that CD38 analyses are technologically stable. Flow cytometry is thus the preferred method to monitor fluctuations in CD38 MFI (CD38 molecules/cell) associated with HIV-disease progression and/or response to ART and has potential for application across instruments and centers. © 2009 Clinical Cytometry Society.
CD3 antigen; CD38 antigen; CD4 antigen; CD8 antigen; antigen expression; article; CD4 lymphocyte count; CD8+ T lymphocyte; controlled study; diagnostic accuracy; diagnostic test; disease course; flow cytometry; human; Human immunodeficiency virus; Human immunodeficiency virus infected patient; Human immunodeficiency virus infection; major clinical study; medical research; patient monitoring; priority journal; quality control; Antigens, CD38; Antigens, CD45; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Disease Progression; Flow Cytometry; HIV Infections; HIV-1; Humans; Quality Control; Reproducibility of Results; T-Lymphocyte Subsets