A sensitive method for detecting proliferation of rare autoantigen- specific human T cells

Stuart I. Mannering, Jessica S. Morris, Kent P. Jensen, Anthony W. Purcell, Margo C. Honeyman, Peter M. Van Endert, Leonard C. Harrison

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Abstract

The ability to measure proliferation of rare antigen-specific T cells among many bystanders is critical for the evaluation of cellular immune function in health and disease. T-cell proliferation in response to antigen has been measured almost exclusively by 3H-thymidine incorporation. This method does not directly identify the phenotype of the proliferating cells and is frequently not sufficiently sensitive to detect rare autoantigen-specific T cells. To overcome these problems, we developed a novel assay for antigen-specific human T-cell proliferation. Peripheral blood mononuclear cells (PBMC) were labelled with the fluorescent dye 5,6-carboxylfluorescein diacetate succinimidyl ester (CFSE) and cells that proliferated in response to antigen, with resultant reduction in CFSE intensity, were measured directly by flow cytometry. This assay was more sensitive than 3H-thymidine incorporation and detected the proliferation of rare antigen-specific CD4 + T cells at 10-fold lower antigen concentrations. It also allowed the phenotype of the proliferating cells to be directly determined. Using the CFSE assay we were able to measure directly the proliferation of human CD4 + T cells from healthy donors in response to the type 1 diabetes autoantigens glutamic acid decarboxylase (GAD) and proinsulin (PI).

Original languageEnglish
Pages (from-to)173-183
Number of pages11
JournalJournal of Immunological Methods
Volume283
Issue number1-2
DOIs
Publication statusPublished - 1 Jan 2003
Externally publishedYes

Keywords

  • Autoimmune disease
  • CFSE
  • Glutamic acid decarboxylase
  • Proinsulin
  • Proliferation
  • Type 1 diabetes

Cite this

Mannering, S. I., Morris, J. S., Jensen, K. P., Purcell, A. W., Honeyman, M. C., Van Endert, P. M., & Harrison, L. C. (2003). A sensitive method for detecting proliferation of rare autoantigen- specific human T cells. Journal of Immunological Methods, 283(1-2), 173-183. https://doi.org/10.1016/j.jim.2003.09.004