title: An adaptation of recombinant vaccinia-based ELISPOT and intracellular cytokine staining for a comparative measurement of cellular immune responses in HIV-1 and HIV-2 infections in West Africa.
abstract: An efficient and quantitative tool for rapid assessment of human immunodeficiency virus (HIV)-induced cellular immune responses is important for resource-limited settings, such as in sub-Saharan Africa. Modifications are required to previously reported methods for evaluating ex-vivo antigen-specific cellular responses based on direct recombinant vaccinia virus (rVV) stimulation of peripheral blood mononuclear cells (PBMCs) by enzyme linked immunosorbent assay (ELISPOT) and by flow cytometry intracellular cytokine assay (ICA). We made such modifications in order to detect specific responses and compared quantitative cellular immune responses in HIV-1 and HIV-2 infected Gambians. The sensitivity of the rVV-based ELISPOT assay was on average 1.25 interferon (IFN)-gamma spot forming cells (SFC) per 50 000 PBMCs specific for either infection, and 5 IFN-gamma-secreting CD8+ T cells/50 000 in the ICA. The level of IFN-gamma SFC detected by ELISPOT and by ICA were correlated (P < 0.02). ICA detected pol-specific responses in 88% and 67% of HIV-1 and HIV-2 subjects, respectively, and gag-specific responses in more than 80% of both infections. Lower proportions of responders were obtained with ELISPOT, for which pol responses were present in 60% of HIV-1 and 46% of HIV-2 infected patients, and gag responses in 55% and 69%, respectively. The assays did not show any significant difference in cellular immune responses between HIV-1 and HIV-2 infected subjects with CD4% >or= 20%. These outcomes are comparable with results obtained using standard techniques and thus this method is a suitable, rapid and less expensive assessment of cellular immunity.
authors.affiliation: Medical Research Council Laboratories, Banjul, The Gambia.
authors.forename: A
authors.initials: A
authors.lastname: Eshofonie
authors.forename: M Shim
authors.initials: MS
authors.lastname: van der Loeff
authors.forename: H
authors.initials: H
authors.lastname: Whittle
authors.forename: A
authors.initials: A
authors.lastname: Jaye
mesh_terms: D000328:Adult; D000368:Aged; D000956:Antigens, Viral / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D015750:Genes, gag; D015751:Genes, pol; D005822:Genetic Vectors; D015658:HIV Infections / Q000276:immunology*; D015497:HIV-1 / Q000235:genetics / Q000276:immunology*; D015498:HIV-2 / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007111:Immunity, Cellular; D007371:Interferon-gamma / Q000096:biosynthesis; D008213:Lymphocyte Activation / Q000276:immunology; D008297:Male; D008875:Middle Aged; D014616:Vaccinia virus / Q000235:genetics / Q000276:immunology
chemical_list: D000956:Antigens, Viral; D007371:Interferon-gamma