Fig. 1 | Mucosal Immunology

Fig. 1

From: Antigen-specific regulatory T-cell responses against aeroantigens and their role in allergy

Fig. 1

Detection tools for antigen-specific CD4+ T-cell analysis and how they reveal different aspects of antigen-specific responses. The cartoon depicts the total repertoire of antigen-specific T cells. Colored cells indicate the T-cell subset(s), which can be detected by the respective detection methods in healthy (upper rows) or allergic donors (lower rows). All available detection methods lead to a certain bias by favoring or missing different T-cell populations. a Long-term (days–weeks) in vitro activation protocols detect proliferating antigen-reactive T cells but modulate the ex vivo phenotype or frequencies. Thus, antigen-reactive T cells may not derive from in vivo antigen-primed memory T cells but instead from naive or cross-reactive memory T cells naturally present human memory pool due to its highly diverse TCR repertoire. b Analysis of T cells expressing particular effector cytokines after short-term stimulation (e.g. ELISPOT, ELISA, and cytokine staining) may be useful to detect functionally important T-cell subsets. However, other subsets, such as naive or anergic T cells as well as Tregs or effector T cells producing other or no cytokines are missed. Cross-reactive memory T cells [see under a)] can also generate false-positive signals. c pMHC-multimers allow in principal the detection of all antigen-specific T-cell subsets, independent of their phenotypic or functional properties. Since they require prior knowledge of antigenic epitopes and MHC alleles, pMHC-multimers may be less suitable for screening of unknown antigen-specificities from complex antigens, e.g. like allergen extracts. Target specificities and affinity may differ between antigen-specific subsets, such as Tcons and Tregs or cross-reactive T cells. This may influence the composition of the T-cell pool detected by pMHC-multimers. d Detection of antigen-reactive T cells based on universal CD4+ T-cell activation markers such CD154 (CD40L) and CD137 (4-1BB) being expressed after only a few hours (5–7 h) of activation allows detection of naive and all antigen-experienced antigen-specific Tcons (CD154+CD137−) and Tregs (CD154−CD137+). This marker combination comprises all antigen-reactive CD4+ T-cell subsets, but per definition misses anergic cells. Importantly, it can be used with any type of antigen, since proteins and naturally processed by autologous APCs, thus providing a global view on the antigen-reactive T-cell response, even for complex antigens