title: Ex vivo analysis of human antigen-specific CD8+ T-cell responses: quality assessment of fluorescent HLA-A2 multimer and interferon-gamma ELISPOT assays for patient immune monitoring.
abstract: The authors developed a standardized approach for immune monitoring of antigen-specific CD8+ T cells within peripheral blood lymphocytes (PBLs) that combines direct ex vivo analysis of Melan-A/MART-1 and influenza-specific CD8+ T cells with HLA-A2/peptide multimers and interferon-gamma ELISPOT assays. Here the authors assessed the quality of results obtained with 180 PBLs from healthy donors and melanoma patients. Reproducibility of the multimer assay was good (average of 15% variation). In the absence of in vivo antigen-specific T-cell responses, physiologic fluctuations of multimer-positive T cells was low, with variation coefficients of 20% for Melan-A and 28% for influenza-specific T cells. In contrast, patients with vaccination-induced T-cell responses had significantly increased T-cell frequencies clearly exceeding physiologic fluctuations. Comparable results were obtained with ELISPOT assays. In conclusion, this approach is well suited to assess T-cell responses as biologic endpoints in clinical vaccine studies.
authors.affiliation: Division of Clinical Onco-Immunology, Ludwig Institute for Cancer Research, Centre Hospitalier Universitaire Vaudois, Lausanne, Switzerland. daniel.speiser@hospvd.ch
authors.forename: Daniel E
authors.initials: DE
authors.lastname: Speiser
authors.forename: Mikaël J
authors.initials: MJ
authors.lastname: Pittet
authors.forename: Philippe
authors.initials: P
authors.lastname: Guillaume
authors.forename: Norbert
authors.initials: N
authors.lastname: Lubenow
authors.forename: Eric
authors.initials: E
authors.lastname: Hoffman
authors.forename: Jean-Charles
authors.initials: JC
authors.lastname: Cerottini
authors.forename: Pedro
authors.initials: P
authors.lastname: Romero
mesh_terms: D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D005434:Flow Cytometry; D005453:Fluorescence; D015789:HLA-A2 Antigen / Q000097:blood* / Q000276:immunology*; D006801:Humans; D007251:Influenza, Human / Q000276:immunology; D007371:Interferon-gamma / Q000097:blood* / Q000276:immunology*; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism; D008545:Melanoma / Q000276:immunology / Q000378:metabolism; D015166:Monitoring, Immunologic / Q000379:methods*; D010455:Peptides / Q000276:immunology; D015203:Reproducibility of Results; D013379:Substrate Specificity; D014611:Vaccination
chemical_list: D015789:HLA-A2 Antigen; D010455:Peptides; D007371:Interferon-gamma