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Fuchs, Y.F.* ; Jainta, G.W.* ; Kühn, D.* ; Wilhelm, C.* ; Weigelt, M.* ; Karasinsky, A.* ; Upadhyaya, B.* ; Ziegler, A.-G. ; Bonifacio, E.

Vagaries of the ELISpot assay: Specific detection of antigen responsive cells requires purified CD8+ T cells and MHC class I expressing antigen presenting cell lines.

Clin. Immunol. 157, 216-225 (2015)
DOI Verlagsversion bestellen
Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.
Quantification of antigen-specific CD8(+) T cells is important for monitoring infection, vaccination, and response to therapy in cancer and immune-mediated diseases. Cytokine enzyme-linked-immunospot (ELISpot) assays are often used for this purpose. We found that substantial spot formation in IFNγ ELISpot assays occurred independently of CD8(+) T cells even when classical MHC class I restricted peptides are used for stimulation. Using fractionated cells and intracellular cytokine staining, the non-CD8(+) T cell IFNγ production was attributed to the CD4(+) T cell fraction. We therefore refined a cell line-based ELISpot assay combining HLA-A*0201 expressing K562 cells for antigen presentation with purified CD8(+) T cells and demonstrated that it specifically detected CD8(+) T cell responses with detection limits comparable to traditional ELISpot assays and dextramer-based quantification. The assay was further adapted to whole antigen responses with antigen (pre-proinsulin)-expressing HLA-A*0201K562 cells. Thus, we revealed and corrected a weak spot of the CD8(+) ELISpot assay.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Schlagwörter Cd8(+) T Cells ; Elispot ; Mhc Class I ; Autoantigen ; Diabetes; Interferon-gamma; Lymphocytes-t; Nod Mice; Peptides; Binding; Identification; Molecules; Recognition; Autoantigens; Autoimmunity
ISSN (print) / ISBN 1521-6616
e-ISSN 1521-7035
Zeitschrift Clinical Immunology
Quellenangaben Band: 157, Heft: 2, Seiten: 216-225 Artikelnummer: , Supplement: ,
Verlag Elsevier
Verlagsort San Diego
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Diabetes Research Type 1 (IDF)
Institute for Pancreatic Beta Cell Research (IPI)