Comparison of Seven Quantitative Assays to Assess Lymphocyte Cell Death during HIV Infection: Measurement of Induced Apoptosis in Anti-Fas-Treated Jurkat Cells and Spontaneous Apoptosis in Peripheral Blood Mononuclear Cells from Children Infected with HIV
- 1 November 1998
- journal article
- research article
- Published by Mary Ann Liebert Inc in AIDS Research and Human Retroviruses
- Vol. 14 (16), 1413-1422
- https://doi.org/10.1089/aid.1998.14.1413
Abstract
The study of apoptosis in relation to various human disease states, particularly HIV infection, has seen a tremendous increase in activity. In this article, values obtained by seven different assays, designed to quantify apoptosis and applicable to the study of HIV infection, are compared in two cell systems: (1) stimulus-induced apoptosis in Jurkat cells treated with anti-Fas antibody and (2) spontaneous apoptosis in PBMCs isolated from HIV-infected children. The methods used included measurement of cells with subdiploid DNA content, labeling of DNA strand breaks by the TUNEL reaction, annexin V surface labeling for the detection of exposed phosphatidylserine, cytoplasmic antigen labeling with the apoptosis-specific antibody Apo 2.7, detection of changes in flow cytometric light-scattering properties, trypan blue dye exclusion by light microscopy, and detection of changes in cellular chromatin by fluorescence microscopy. These methods produced well-correlated values in the Jurkat system, whereas the same set of methods produced more discrepant values in the PBMC analyses, especially in those patients with low CD4 counts. Specifically, our results showed that the trypan blue test was unacceptable for quantification of apoptosis during HIV infection, whereas TUNEL, of all the methods tested, showed excellent overall correlation in both cell systems, was highly specific, and matched microscopic observation of the cells. Although many of the methods were suited to the study of a homogeneous cell line, caution must be exercised when examining cell death in a heterogeneous cell mixture from an HIV-infected individual.Keywords
This publication has 37 references indexed in Scilit:
- Supravital exposure to propidium iodide identifies apoptotic cells in the absence of nucleosomal DNA fragmentationCytometry, 1996
- Use of CD45 fluorescence and side‐scatter characteristics for gating lymphocytes when using the whole blood lysis procedure and flow cytometryCytometry, 1996
- Discrimination of apoptotic thymocytes by forward light scatterCytometry, 1995
- Further characterisation of the in situ terminal deoxynucleotidyl transferase (TdT) assay for the flow cytometric analysis of apoptosis in drug resistant and drug sensitive leukaemic cellsCytometry, 1995
- Apoptosis without decrease of cell DNA contentFEBS Letters, 1995
- Single‐step procedure for labeling DNA strand breaks with fluorescein‐ or Bodipy‐conjugated deoxynucleotides: Detection of apoptosis and bromodeoxyuridine incorporationCytometry, 1995
- Autocrine T-cell suicide mediated by APO-1/(Fas/CD95)Nature, 1995
- Use of a Flow Cytometric Assay to Quantitate Apoptosis in Human LymphocytesClinical Immunology and Immunopathology, 1994
- Programmed Death of T Cells in HIV-1 InfectionScience, 1992
- Activation-induced death by apoptosis in CD4+ T cells from human immunodeficiency virus-infected asymptomatic individuals.The Journal of Experimental Medicine, 1992