ELISPOT and intracellular cytokine staining: Novel assays for quantifying T cell responses in the chicken

M. P. Ariaans, P. M. van de Haar, J. W. Lowenthal, W. van Eden, E. J. Hensen, L. Vervelde*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


The measurement of T cell responses in chickens, not only for quantitative aspects but also for the qualitative nature of the responses, becomes increasingly important. However, there are very few assays available to measure T cell function. Therefore, we have developed enzyme-linked immunosorbent spot assay (ELISPOT) and an intracellular cytokine staining (ICCS) assay. ELISPOT assay for the detection of chicken interferon-gamma (ChIFN-gamma) production was set up and shown to be reproducible for both polyclonal and antigen-specific stimuli such as Newcastle disease virus (NDV). However, the ELISPOT assay lacks the ability to identify individual cytokine-producing cells. Separation of CD4(+) and CD8(+) T cell populations gave additional information, but appeared to have the disadvantage of a loss of cell interactions during stimulation. In a further refinement, individual cells were identifiable by ICCS, which gives the possibility to characterize for multiple characteristics, such as cytokine production and phenotype of the cell. Using ICCS, ChIFN-gamma production was evaluated. Although cells were detected at only low frequencies, polyclonal stimulation of peripheral, blood mononuclear cell (PBMC) or spleen cells resulted in a significant increase in ChIFN-gamma production by CD4(+) and CD8(+) cells. (C) 2008 Elsevier Ltd. All rights reserved.

Original languageEnglish
Pages (from-to)1398-1404
Number of pages7
JournalDevelopmental and Comparative Immunology
Issue number11
Publication statusPublished - 2008


  • chicken
  • interferon-gamma
  • T cell assay

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