Optimization of a Method to Detect Autoantigen-Specific T-Cell Responses in Type 1 Diabetes
Open Access
- 7 December 2020
- journal article
- research article
- Published by Frontiers Media SA in Frontiers in Immunology
Abstract
The development of tolerizing therapies aiming to inactivate autoreactive effector T-cells is a promising therapeutic approach to control undesired autoimmune responses in human diseases such as Type 1 Diabetes (T1D). A critical issue is a lack of sensitive and reproducible methods to analyze antigen-specific T-cell responses, despite various attempts. We refined a proliferation assay using the fluorescent dye 5,6-carboxylfluorescein diacetate succinimidyl ester (CFSE) to detect responding T-cells, highlighting the fundamental issues to be taken into consideration to monitor antigen-specific responses in patients with T1D. The critical elements that maximize detection of antigen-specific responses in T1D are reduction of blood storage time, standardization of gating parameters, titration of CFSE concentration, selecting the optimal CFSE staining duration and the duration of T-cell stimulation, and freezing in medium containing human serum. Optimization of these elements enables robust, reproducible application to longitudinal cohort studies or clinical trial samples in which antigen-specific T-cell responses are relevant, and adaptation to other autoimmune diseases.Keywords
This publication has 30 references indexed in Scilit:
- Serum-free freezing media support high cell quality and excellent ELISPOT assay performance across a wide variety of different assay protocolsCancer Immunology, Immunotherapy, 2012
- Structural basis for the killing of human beta cells by CD8+ T cells in type 1 diabetesNature Immunology, 2012
- Standardized Serum-Free Cryomedia Maintain Peripheral Blood Mononuclear Cell Viability, Recovery, and Antigen-Specific T-Cell Response Compared to Fetal Calf Serum-Based MediumBiopreservation and Biobanking, 2011
- Current approaches to measuring human islet-antigen specific T cell function in type 1 diabetesClinical and Experimental Immunology, 2010
- Recurrence of Type 1 Diabetes After Simultaneous Pancreas-Kidney Transplantation, Despite Immunosuppression, Is Associated With Autoantibodies and Pathogenic Autoreactive CD4 T-CellsDiabetes, 2010
- Standardization and optimization of multiparameter intracellular cytokine stainingCytometry Part A, 2008
- Responses against islet antigens in NOD mice are prevented by tolerance to proinsulin but not IGRPJCI Insight, 2006
- Analysis of T-Cell Assays to Measure Autoimmune Responses in Subjects With Type 1 DiabetesDiabetes, 2006
- The insulin A-chain epitope recognized by human T cells is posttranslationally modifiedThe Journal of Experimental Medicine, 2005
- Functional avidity directs T-cell fate in autoreactive CD4+ T cellsBlood, 2005