Abstract
We used a novel approach for the direct ex vivo identification and characterization of T cell epitopes based on the screening of peptide spot libraries with freshly isolated splenocytes in a sensitive enzyme-linked immunospot (ELISPOT) assay. This technique was applied for the analysis of splenocytes from Listeria monocytogenes-infected BALB/c and C57BL/6 mice. The screening of peptide spot libraries covering the whole listeriolysin O and p60 of L. monocytogenes confirmed all known CD4 and CD8 T cell epitopes of these proteins and additionally revealed six new H-2(d) and six new H-2(b)-restricted T cell epitopes. New epitopes were categorized into CD4 and CD8 T cell epitopes by ex vivo ELISPOT analysis with separated T cell populations. The quantitative analysis of cells reactive with these CD4 and CD8 T cell epitopes revealed the existence of dominant and subdominant CD4 and CD8 T cell populations during L. monocytogenes infection. As a consequence of these data we suggest that ELISPOT-based screening of peptide spot libraries could be a general approach for the rapid identification and characterization of pathogen-specific T cell populations during various infectious diseases.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Bacterial Proteins / analysis
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Bacterial Proteins / immunology
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Bacterial Toxins*
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CD4-Positive T-Lymphocytes / immunology*
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CD4-Positive T-Lymphocytes / metabolism
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CD4-Positive T-Lymphocytes / microbiology*
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CD8-Positive T-Lymphocytes / immunology*
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CD8-Positive T-Lymphocytes / metabolism
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CD8-Positive T-Lymphocytes / microbiology*
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Cell Line
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Enzyme-Linked Immunosorbent Assay / methods
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Epitope Mapping / methods*
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Epitopes, T-Lymphocyte / analysis*
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Epitopes, T-Lymphocyte / immunology
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Female
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Heat-Shock Proteins / analysis
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Heat-Shock Proteins / immunology
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Hemolysin Proteins
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Histocompatibility Antigens Class I / analysis
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Histocompatibility Antigens Class I / immunology
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Histocompatibility Antigens Class II / analysis
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Histocompatibility Antigens Class II / immunology
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Immunodominant Epitopes / analysis*
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Immunodominant Epitopes / immunology
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Immunologic Memory
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Immunophenotyping
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Listeria monocytogenes / immunology*
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Lymphocyte Activation
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Mice
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Mice, Inbred BALB C
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Mice, Inbred C57BL
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Peptide Library
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T-Lymphocyte Subsets / immunology
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T-Lymphocyte Subsets / metabolism
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T-Lymphocyte Subsets / microbiology
Substances
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60 kDa protein, Listeria monocytogenes
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Bacterial Proteins
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Bacterial Toxins
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Epitopes, T-Lymphocyte
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Heat-Shock Proteins
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Hemolysin Proteins
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Histocompatibility Antigens Class I
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Histocompatibility Antigens Class II
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Immunodominant Epitopes
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Peptide Library
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hlyA protein, Listeria monocytogenes