Regulatory T cells play a critical role in down regulating immune responses. Although CD25 positivity has been used as a marker of these cells, other markers, such as surface TGF-beta may be more specific and may yield greater insights into regulatory T cell function. The goal of this study was to characterize the amplification capacity of phycobilisomes and employ them in the detection of cell surface TGF-beta.

To quantify amplification, streptavidin conjugated phycobilisomes and allophycocyanin (APC) were compared. Surface TGF-beta was examined on activated CD25+ and CD25- CD4 T cells by staining with TGF-beta biotin streptavidin-phycobilisome.

Use of phycobilisomes increased fluorescent intensity 100-1000 fold relative to conventional APC conjugates. Despite this amplification, nonspecific staining was minimally increased. Using phycobilisome amplification, we found that in CD3 activated CD4 T cells, surface TGF-beta was found predominantly in the CD4+, CD25+ subset (CD25+ vs. CD25− : 62% vs. 35%, p=0.007, n=3). Surprisingly, surface TGF-beta was detectable even in unactivated T cells and was similarly found to be higher in the CD4+CD25+ subset (CD25+ vs. CD25− : 29% vs. 8%, p=0.002 n=3).

Phycobilisomes provide a unique capability for staining low density cell surface molecules such as surface TGF-beta. Surface TGFbeta expression may be a useful phenotypic marker for regulatory T cells. The detection of CD25 negative, cell surface TGF-beta expressing T cells may indicate the presence of substantial regulatory T cell activity outside of the classic CD25+ subset.