title: Use of an in vivo FTA assay to assess the magnitude, functional avidity and epitope variant cross-reactivity of T cell responses following HIV-1 recombinant poxvirus vaccination.
abstract: Qualitative characteristics of cytotoxic CD8+ T cells (CTLs) are important in measuring the effectiveness of CTLs in controlling HIV-1 infections. Indeed, in recent studies patients who are naturally resistant to HIV-1 infections have been shown to possess CTLs that are of high functional avidity and have a high capacity to recognize HIV epitope variants, when compared to HIV-1 infection progressors. When developing efficacious vaccines, assays that can effectively measure CTL quality specifically in vivo are becoming increasingly important. Here we report the use of a recently developed high-throughput multi-parameter technique, known as the fluorescent target array (FTA) assay, to simultaneously measure CTL killing magnitude, functional avidity and epitope variant cross-reactivity in real time in vivo. In the current study we have applied the FTA assay as a screening tool to assess a large cohort of over 20 different HIV-1 poxvirus vaccination strategies in mice. This screen revealed that heterologous poxvirus prime-boost vaccination regimes (i.e., recombinant fowlpox (FPV)-HIV prime followed by a recombinant vaccinia virus (VV)-HIV booster) were the most effective in generating high quality CTL responses in vivo. In conclusion, we have demonstrated how the FTA assay can be utilized as a cost effective screening tool (by reducing the required number of animals by >100 fold), to evaluate a large range of HIV-1 vaccination strategies in terms of CTL avidity and variant cross-reactivity in an in vivo setting.
authors.affiliation: Molecular Mucosal Vaccine Immunology Group, The John Curtin School of Medical Research, The Australian National University, Canberra, Australia.
authors.forename: Danushka K
authors.initials: DK
authors.lastname: Wijesundara
authors.affiliation: Molecular Mucosal Vaccine Immunology Group, The John Curtin School of Medical Research, The Australian National University, Canberra, Australia.
authors.forename: Charani
authors.initials: C
authors.lastname: Ranasinghe
authors.affiliation: Molecular Mucosal Vaccine Immunology Group, The John Curtin School of Medical Research, The Australian National University, Canberra, Australia.
authors.forename: Ronald J
authors.initials: RJ
authors.lastname: Jackson
authors.affiliation: Alternatives to Animals through Bioinformatics Group, Dept Genome Biology, The John Curtin School of Medical Research, The Australian National University, Canberra, Australia.
authors.forename: Brett A
authors.initials: BA
authors.lastname: Lidbury
authors.affiliation: Cancer and Vascular Biology Group, Dept Immunology, The John Curtin School of Medical Research, The Australian National University, Canberra, Australia.
authors.forename: Christopher R
authors.initials: CR
authors.lastname: Parish
authors.affiliation: Cancer and Vascular Biology Group, Dept Immunology, The John Curtin School of Medical Research, The Australian National University, Canberra, Australia.
authors.forename: Benjamin J C
authors.initials: BJ
authors.lastname: Quah
mesh_terms: D016915:AIDS Vaccines / Q000008:administration & dosage / Q000276:immunology*; D000818:Animals; D003429:Cross Reactions; D000939:Epitopes / Q000276:immunology*; D005453:Fluorescence; D015658:HIV Infections / Q000276:immunology / Q000517:prevention & control*; D015497:HIV-1 / Q000276:immunology*; D057166:High-Throughput Screening Assays / Q000379:methods; D006801:Humans; D007117:Immunization, Secondary; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D046228:Microarray Analysis / Q000379:methods; D011212:Poxviridae / Q000276:immunology*; D011213:Poxviridae Infections / Q000276:immunology / Q000517:prevention & control*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D014611:Vaccination; D019444:Vaccines, DNA / Q000008:administration & dosage / Q000276:immunology*; D014616:Vaccinia virus / Q000276:immunology
chemical_list: D016915:AIDS Vaccines; D000939:Epitopes; D019444:Vaccines, DNA