title: Improving the sensitivity of the ELISPOT analyses of antigen-specific cellular immune responses in rhesus macaques.
abstract: The close similarities in hematopoietic and immune systems of humans and rhesus macaques (Macaca mulatta) make them the desired nonhuman primate animal model for developing vaccines against infectious diseases relevant to humans. The best example is the simian immunodeficiency virus infection in macaques as a model for AIDS, resulting from infection of humans by HIV-1. Vaccine efficacy against viruses depends on priming cell-mediated immunity by the use of sensitive assays that can accurately detect even small levels of antigen-specific T-cell responses that may otherwise be missed easily. With this in mind, we developed the dendritic cell enzyme-linked immunospot (DC-ELISPOT) protocol by incorporating antigen-pulsed DCs to stimulate lymphocytes, as opposed to the conventional ELISPOT assay, which cultures mixtures of various low-level populations of indigent antigen-presenting cells and responding lymphocytes with antigens. In rhesus macaques immunized with an HIV envelope peptide cocktail vaccine, the DC-ELISPOT protocol enabled more accurate enumeration of the cellular immune responses, as antigen-specific interferon-gamma-producing cells, with up to 18-fold increase in detection sensitivity compared with conventional ELISPOT and elimination of false negative results. The increased sensitivity of DC-ELISOT protocol is further validated in tests determining recall antigen-specific responses in human volunteers after tuberculin skin testing.
authors.affiliation: Department of Immunology, University of Texas M.D. Anderson Cancer Center, Houston, TX, USA.
authors.forename: K Jagannadha
authors.initials: KJ
authors.lastname: Sastry
authors.forename: Pramod N
authors.initials: PN
authors.lastname: Nehete
authors.forename: Bharti
authors.initials: B
authors.lastname: Nehete
mesh_terms: D000818:Animals; D017951:Antigen Presentation; D000941:Antigens; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D003713:Dendritic Cells / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000706:statistics & numerical data*; D015488:HIV Antigens / Q000008:administration & dosage; D006801:Humans; D007111:Immunity, Cellular; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000032:analysis / Q000096:biosynthesis; D008253:Macaca mulatta; D012680:Sensitivity and Specificity
chemical_list: D000941:Antigens; D015488:HIV Antigens; D007371:Interferon-gamma