Detection of low-frequency human antigen-specific CD4+ T cells using MHC class II multimer bead sorting and immunoscope analysis

Authors

  • Fabrice Lemaître,

    Corresponding author
    1. Unité de Recherche et d'Expertise Immunité anti-virale, Biothérapie et Vaccins, INSERM U277, Institut Pasteur, Paris, France
    • INSERM U277, Unité de Recherche et d'Expertise Immunité anti-virale, Biothérapie et Vaccins, Institut Pasteur, 25 rue du Docteur Roux, F-75724 Paris Cedex 15, France, Fax: +33-1-4568-8548
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    • The first two authors contributed equally to this work.

  • Manuelle Viguier,

    1. Unité de Recherche et d'Expertise Immunité anti-virale, Biothérapie et Vaccins, INSERM U277, Institut Pasteur, Paris, France
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    • The first two authors contributed equally to this work.

  • Min-Sun Cho,

    1. Unité de Recherche et d'Expertise Immunité anti-virale, Biothérapie et Vaccins, INSERM U277, Institut Pasteur, Paris, France
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  • Jean-Marie Fourneau,

    1. INSERM U580, Institut Necker, Paris, France
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  • Bernard Maillère,

    1. Département d'ingénierie et d'étude des protéines, CEA-Saclay, Gif-sur-Yvette, France
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  • Philippe Kourilsky,

    1. Unité de Recherche et d'Expertise Immunité anti-virale, Biothérapie et Vaccins, INSERM U277, Institut Pasteur, Paris, France
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  • Peter M. van Endert,

    1. INSERM U580, Institut Necker, Paris, France
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  • Laurent Ferradini

    1. Unité de Recherche et d'Expertise Immunité anti-virale, Biothérapie et Vaccins, INSERM U277, Institut Pasteur, Paris, France
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Abstract

MHC class II tetramers are attractive tools to study antigen-specific CD4+ T cell responses in various clinical situations in humans. HLA-DRA1*0101/DRB1*0401 MHC class II heterodimers were produced as empty molecules using the Drosophila melanogaster expression system. Peptide binding experiments revealed that these molecules could be loaded efficiently with appropriate MHC class II tumor epitopes. Interestingly, MHC class II tetramer staining was influenced by modifications in membrane lipid rafts, and could in itself induce activation changes of stained CD4+ T cells at 37°C. In order to increase the threshold of detection of poorly represented peripheral antigen-specific CD4+ T cells, we combined cell sorting using MHC class II multimer beads together with TCR analysis using the immunoscope technology. This strategy greatly increased the sensitivity of detection of specific CD4+ T cells to frequencies as low as 4×10–6 among peripheral blood mononuclear cells. Such a combined approach may have promising applications in the immunomonitoring of patients under vaccination protocols to tightly follow induced antigen-specific CD4+ T cells expressing previously identified TCR.

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