Antigen-specific Treg and Teff can be detected by measuring down-regulation of surface.

<p><i>CD3</i>. PBMC from Patient 113 were cultured with pooled NY-ESO-1 18mer peptides as described in <i>Methods</i>, followed by re-stimulation with the indicated individual peptides during overnight culture to allow CD3 down-regulation. Cells were stained with antibodies to CD4, CD25, FoxP3 and CD3 and analyzed by flow cytometry. (<b><i>A</i></b>): an example of the staining patterns observed, illustrating the gating of Treg and Teff (on gated viable CD4⁺ T cells) and the down-regulation of CD3 within each population. (<b><i>B</i></b>–<b><i>C</i></b>): A summary of the responses detected within the Treg (B) and Teff (C) populations. Dotted lines indicate the baseline level of CD3-low cells (nil peptide condition).