Ex vivo T cell-based HIV suppression assay to evaluate HIV-specific CD8+ T-cell responses.
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.
Unité de Régulation des Infections Rétrovirales, Institut Pasteur, Paris, France. asier.saez-cirion@pasteur.fr
Asier
A
Sáez-Cirión
So Youn
SY
Shin
Pierre
P
Versmisse
Françoise
F
Barré-Sinoussi
Gianfranco
G
Pancino
D016915:AIDS Vaccines / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology / Q000821:virology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology* / Q000821:virology*; D004797:Enzyme-Linked Immunosorbent Assay; D016655:HIV Core Protein p24 / Q000378:metabolism; D015658:HIV Infections / Q000276:immunology / Q000517:prevention & control / Q000821:virology; D020276:HIV Long-Term Survivors; D015497:HIV-1 / Q000276:immunology* / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D014779:Virus Replication / Q000276:immunology; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000378:metabolism
D016915:AIDS Vaccines; D016655:HIV Core Protein p24; D054301:gag Gene Products, Human Immunodeficiency Virus; C514799:p24 protein, Human Immunodeficiency Virus Type 1