about 1–2% of Treg can be measured in whole blood. The additional measurement of cellular expression of FOXP3 protein allowed a more specific analysis of Treg cells (CD4+CD25+FOXP3+ cells). However, FOXP3 is also transiently expressed in activated human effector T cells, thus complicating a correct Treg analysis using CD4, CD25 and FOXP3 as markers in humans. Therefore, the gold standard surface marker combination to defined Tregs within unactivated CD3+CD4+ T cells is high CD25 expression combined with the absent or low-level expression of the surface protein CD127 (IL-7RA). If viable cells are not required then the addition of FOXP3 to the CD25 and CD127 combination will provide further stringency. Several additional markers have been described, e.g., high levels of CTLA-4 (cytotoxic T-lymphocyte associated molecule-4) and GITR (glucocorticoid-induced TNF receptor) are also expressed on regulatory T cells, however the functional significance of this expression remains to be defined. There is a great interest in identifying cell surface markers that are uniquely and specifically expressed on all FOXP3-expressing regulatory T cells. However, to date no such molecule has been identified. The identification of Tregs following cell activation is challenging as conventional T cells will express CD25, transiently express FOXP3 and lose CD127 expression upon activation. It has been shown that Tregs can be detected using an activation-induced marker assay by expression of CD39 in combination with co-expression of CD25 and OX40(CD134) which define antigen-specific cells following 24-48h stimulation with antigen. In addition to the search for novel protein markers, a different method to analyze and monitor Treg cells more accurately has been described in the literature. This method is based on DNA methylation analysis. Only in Treg cells, but not in any other cell type, including activated effector T cells, a certain region within the gene (TSDR, Treg-specific-demethylated region) is found demethylated, which allows to monitor Treg cells through a PCR reaction or other DNA-based analysis methods. Interplay between the Th17 cells and regulatory T cells are important in many diseases like respiratory diseases. Recent evidence suggests that mast cells may be important mediators of Treg-dependent peripheral tolerance. Epitopes Regulatory T cell epitopes ('Tregitopes') were discovered in 2008 and consist of linear sequences of amino acids contained within monoclonal antibodies and immunoglobulin G (IgG). Since their discovery, evidence has indicated Tregitopes may be crucial to the activation of natural regulatory T cells. Potential applications of regulatory T cell epitopes have been hypothesised: