title: Ultra-sensitive class I tetramer analysis reveals previously undetectable populations of antiviral CD8+ T cells.
abstract: A major breakthrough in cellular immunology has been the development of HLA class I tetramers to analyze CD8(+) T cell responses. However, in many situations, including persistent virus infection, specific T cell responses are rarely detected using this technology. This raises the question of whether such responses are 'deleted' (or 'exhausted') or present below the conventional detection limit for class I tetramer staining. In particular, persistent hepatitis C virus (HCV) infection is characterized by very weak or apparently absent specific CD8(+) T cell responses, even though they are readily detectable in acute disease. Therefore, we assessed the use of anti-PE-labeled magnetic beads to enrich tetramer-positive HCV-specific T cells and identify previously undetectable populations. Using the enrichment technique, HCV-specific T cells could be detected in the majority of infected individuals, whereas these responses were not detected using conventional tetramer staining (8/15 vs. 1/15; p=0.01). Magnetic enrichment could reliably detect very rare HCV-specific responses at frequencies of >0.0011% of CD8(+) T cells (approximately 1/million PBMC), and phenotypic analysis of these rare populations was possible. Therefore, this direct ex vivo technique revealed the persistence of very low frequencies of virus-specific CD8(+) T cells during chronic virus infection and is readily transferable to the study of other viral, self- or tumor-specific T cells.
authors.affiliation: Peter Medawar Building for Pathogen Research, Nuffield Department of Clinical Medicine, University of Oxford, GB.
authors.forename: Eleanor
authors.initials: E
authors.lastname: Barnes
authors.forename: Scott M
authors.initials: SM
authors.lastname: Ward
authors.forename: Victoria O
authors.initials: VO
authors.lastname: Kasprowicz
authors.forename: Geoffrey
authors.initials: G
authors.lastname: Dusheiko
authors.forename: Paul
authors.initials: P
authors.lastname: Klenerman
authors.forename: Michaela
authors.initials: M
authors.lastname: Lucas
mesh_terms: D000951:Antigens, Neoplasm; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology* / Q000821:virology; D015789:HLA-A2 Antigen / Q000276:immunology*; D016174:Hepacivirus / Q000276:immunology*; D018936:Hepatitis C Antigens / Q000276:immunology / Q000302:isolation & purification; D019698:Hepatitis C, Chronic / Q000276:immunology* / Q000821:virology; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods; D058965:MART-1 Antigen; D009363:Neoplasm Proteins / Q000276:immunology; D010799:Phycoerythrin / Q000276:immunology
chemical_list: D000951:Antigens, Neoplasm; D015789:HLA-A2 Antigen; D018936:Hepatitis C Antigens; D058965:MART-1 Antigen; C548667:MLANA protein, human; D009363:Neoplasm Proteins; D010799:Phycoerythrin