Ex vivo T cell–based HIV suppression assay to evaluate HIV-specific CD8+ T-cell responses
Open Access
- 13 May 2010
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature Protocols
- Vol. 5 (6), 1033-1041
- https://doi.org/10.1038/nprot.2010.73
Abstract
To advance T cell-based HIV vaccine development, it is necessary to evaluate the immune correlates of a protective CD8(+) T-cell response. We have developed an assay that assesses the capacity ex vivo of HIV-specific CD8(+) T cells to suppress HIV-1 infection of autologous CD4(+) T cells. this assay directly reflects the ultimate effector function of CD8(+) T cells, the elimination of infected cells, and accurately differentiates the effective CD8(+) T-cell response in spontaneous HIV controllers from ineffective responses in other patients. In this article, we describe all the steps from cell purification to assessment of viral replication by HIV-p24 ELISA and analysis, along with conditions for cell culturing, and how to choose the viral infectious dose that gives the most reliable results. We also depict the conditions of a rapid assay on the basis of flow cytometry analysis of intracellular HIV-Gag products. these procedures take 14-17 d when the p24 ELISA assay is used, or 6 d with the intracellular Gag assay.Keywords
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