Detection of autoreactive T cells in type 1 diabetes using coded autoantigens and an immunoglobulin-free cytokine ELISPOT assay: report from the fourth immunology of diabetes society T cell workshop.
The "gold standard" for evaluation of immunoassays is blinded testing, using coded samples and antigens. Although assays for autoreactive T cells are no exception to this rule, it is nonetheless rarely applied in this context. To facilitate such investigations, we coded a panel of 10 peptides representing T cell epitopes reported to be of relevance to islet autoimmunity. These were deployed in a novel cytokine ELISPOT assay, in which the use of immunoglobulin-free medium reduces background reactivity and thus potentially enhances the specificity and sensitivity of detection of autoreactive T cells. Significant IFN-gamma production against GAD65 (554-575), insulin (B9-23), and IA-2 (709-736) peptides were observed in type 1 diabetic patients, whereas no significant changes from background were detected in healthy controls. These results confirm the utility of the blinded performance of T cell assays as the most robust platform for assessing technologies to T cell autoreactivity.
Nagata
Masao
M
Department of Immunobiology, Guy's, King's & St Thomas' School of Medicine, 2nd Floor, New Guy's House, Guy's Hospital, London SE1 9RT, UK.
Kotani
Reiko
R
Moriyama
Hiroaki
H
Yokono
Koichi
K
Roep
Bart O
BO
Peakman
Mark
M
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001324:Autoantigens / Q000276:immunology*; D016022:Case-Control Studies; D002478:Cells, Cultured; D018920:Coculture Techniques; D016207:Cytokines / Q000097:blood* / Q000378:metabolism; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology / Q000378:metabolism*; D005240:Feasibility Studies; D005260:Female; D005968:Glutamate Decarboxylase / Q000494:pharmacology; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D007328:Insulin / Q000494:pharmacology; D007371:Interferon-gamma / Q000096:biosynthesis / Q000276:immunology; D007564:Japan; D008297:Male; D008565:Membrane Proteins / Q000276:immunology; D008875:Middle Aged; D010446:Peptide Fragments / Q000494:pharmacology; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000276:immunology; D054635:Receptor-Like Protein Tyrosine Phosphatases, Class 8; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012680:Sensitivity and Specificity; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology*
D001324:Autoantigens; D016207:Cytokines; D007328:Insulin; D008565:Membrane Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D007371:Interferon-gamma; C516862:PTPRN protein, human; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; D054635:Receptor-Like Protein Tyrosine Phosphatases, Class 8; D005968:Glutamate Decarboxylase