Comparison of islet autoantigen-specific T cell response detected by direct enzyme-linked immunospot (ELISPOT) assay and accelerated co-cultured dendritic cells (acDCs) assay

Abstract

Objective: To investigate the effect of enzyme-linked immunospot (ELISPOT) on accelerated co-cultured dendritic cells (acDCs) and direct detection of islet full-length antigen-specific T cell response in peripheral blood of patients with type 1 diabetes mellitus (T1DM). Methods: Sixteen patients with T1DM[9 males, 7 females, mean age(28.5±9.4)years] and 12 age-and sex-matched healthy controls were selected in the Department of Metabolism and Endocrinology, the Second Xiangya Hospital between March 2012 and August 2014. The numbers of IFN-γ secreting CD4(+)T cells responding to glutamic acid decarboxylase 65 (GAD(65)), C-peptide (CP) and insulin (INS) were detected by ELISPOT-acDCs and ELISPOT-direct assays, respectively. The positive rate of islet autoantigen and associated antigen reactive T cells under different detection assays were compared. Results: The positive rate for GAD(65), INS, and CP antigen reactive T cells detected by ELISPOT-acDCs was 1/16, 6/16 and 4/16, respectively, and T cells positive for INS in T1DM patients were higher than that in the controls (0/12) (P=0.024). Combining GAD(65), CP and INS-ELISPOT-acDCs detection, the positive rate for CD4(+) T cells in T1DM patients was higher than that in the controls (9/16 vs 1/12, P=0.016). The positive rate for GAD(65), INS, and CP antigen reactive T cells detected by ELISPOT-direct detection was 2/16, 1/16 and 7/16, respectively, and T cells positive for CP was higher than that in the controls (1/12), but the difference was not statistically significant (P=0.088). Likewise, the positive rate for CD4(+) T cells was higher in T1DM patients than that in the controls by combined GAD(65), CP and INS-ELISPOT-direct detection (8/16 vs 1/12, P=0.039). Compared with the ELISPOT-direct assay, the positive rate of INS antigen specific T cell response detected by ELISPOT-acDCs was higher (P=0.041). No statistical differences of other antigens were found between the two groups (all P>0.05). Conclusions: Both multiple islet antigens-combined CD4(+)-ELISPOT-acDCs and direct assays could provide diagnostic value of cellular immunology for T1DM patients. The ELISPOT-acDCs assay is superior to the ELISPOT-direct assay in the detection of INS antigen-specific T cell response.