• Open Access

Ex-vivo whole blood secretion of interferon (IFN)-γ and IFN-γ-inducible protein-10 measured by enzyme-linked immunosorbent assay are as sensitive as IFN-γ enzyme-linked immunospot for the detection of gluten-reactive T cells in human leucocyte antigen (HLA)-DQ2·5+-associated coeliac disease

Authors

  • N. Ontiveros,

    1. The Walter and Eliza Hall Institute of Medical Research, Parkville, Vic., Australia
    2. Department of Medical Biology, The University of Melbourne, Parkville, Vic., Australia
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  • J. A. Tye-Din,

    1. The Walter and Eliza Hall Institute of Medical Research, Parkville, Vic., Australia
    2. Department of Medical Biology, The University of Melbourne, Parkville, Vic., Australia
    3. Department of Gastroenterology, The Royal Melbourne Hospital, Parkville, Vic., Australia
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  • M. Y. Hardy,

    1. The Walter and Eliza Hall Institute of Medical Research, Parkville, Vic., Australia
    2. Department of Medical Biology, The University of Melbourne, Parkville, Vic., Australia
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  • R. P. Anderson

    Corresponding author
    1. The Walter and Eliza Hall Institute of Medical Research, Parkville, Vic., Australia
    2. Department of Medical Biology, The University of Melbourne, Parkville, Vic., Australia
    3. ImmusanT, Inc., Cambridge, MA, USA
    • Correspondence: R. P. Anderson, ImmusanT, Inc., One Kendall Square, Building 200, LL, Suite 4, Cambridge, MA 02139, USA.

      E-mail: bob@immusant.com

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Summary

T cell cytokine release assays are used to diagnose infectious diseases, but not autoimmune or allergic disease. Coeliac disease (CD) is a common T cell-mediated disease diagnosed by the presence of gluten-dependent intestinal inflammation and serology. Many patients cannot be diagnosed with CD because they reduce dietary gluten before medical workup. Oral gluten challenge in CD patients treated with gluten-free diet (GFD) mobilizes gluten-reactive T cells measurable by interferon (IFN)-γ enzyme-linked immunospot (ELISPOT) or major histocompatibility complex (MHC) class II tetramers. Immunodominant peptides are quite consistent in the 90% of patients who possess HLA-DQ2·5. We aimed to develop whole blood assays to detect gluten-specific T cells. Blood was collected before and after gluten challenge from GFD donors confirmed to have CD (n = 27, all HLA-DQ2·5+), GFD donors confirmed not to have CD (n = 6 HLA-DQ2·5+, 11 HLA-DQ2·5) and donors with CD not following GFD (n = 4, all HLA-DQ2·5+). Plasma IFN-γ and IFN-γ inducible protein-10 (IP-10) were measured by enzyme-linked immunosorbent assay (ELISA) after whole blood incubation with peptides or gliadin, and correlated with IFN-γ ELISPOT. No T cell assay could distinguish between CD patients and controls prior to gluten challenge, but after gluten challenge the whole blood IFN-γ ELISA and the ELISPOT were both 85% sensitive and 100% specific for HLA-DQ2·5+ CD patients; the whole blood IP-10 ELISA was 94% sensitive and 100% specific. We conclude that whole blood cytokine release assays are sensitive and specific for detection of gluten-reactive T cells in CD; further clinical studies addressing the utility of these tests in patients with an uncertain diagnosis of CD is warranted.

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