A dual color ELISPOT method for the simultaneous detection of IL-2 and IFN-γ HIV-specific immune responses

Salix Boulet, Michel L. Ndongala, Yoav Peretz, Marie Pierre Boisvert, Mohamed Rachid Boulassel, Cecile Tremblay, Jean Pierre Routy, Rafick P. Sekaly, Nicole F. Bernard

Research output: Contribution to journalArticlepeer-review

39 Scopus citations


The single color IFN-γ ELISPOT assay has become a standard for assessing HIV-specific immune responses in HIV-infected subjects. However, recent data suggests that single cytokine detection for immune monitoring of HIV-infected individuals may not be sufficient to fully describe virus-specific immune responses. Here, we have designed and validated a dual color ELISPOT assay capable of detecting both IL-2 and IFN-γ secreting cells simultaneously in response to HIV antigens. We found that a cell input number of 200,000 cells/well provided a good balance between limited availability of cells due to blood volume restrictions and ability to detect all cytokine secretion patterns. The simultaneous detection of IL-2 and IFN-γ resulted in a decreased magnitude of IFN-γ but not IL-2 responses. Measures of intra- and inter-assay variability for the dual color ELISPOT assay were comparable to that seen for single cytokine ELISPOT assay with coefficients of variation below 20% for IL-2, IFN-γ and dual secretion. Although CD8+ T cells mediated most HIV-specific responses in infected subjects, CD4+ T cells mediated responses to HIV were also detected. Features of this assay such as high throughput, cell number requirement and cytokine choice should make this assay a valuable tool for screening for HIV-specific immune responses in several clinically relevant settings.

Original languageEnglish (US)
Pages (from-to)18-29
Number of pages12
JournalJournal of Immunological Methods
Issue number1-2
StatePublished - Mar 30 2007
Externally publishedYes


  • CD4+
  • CD8+
  • Cellular immune responses
  • HIV
  • IL-2
  • Interferon-gamma
  • T cells

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology


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