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In this study, by using of near-field scanning optical microscopy (NSOM)/immune-labeling quantum dot (QD)-based dual-color imaging system, we achieved the direct visualization of nanoscale profiles for distribution and organization of CD4 and CD25 molecules in T cells. A novel and interesting finding was that though CD25 clustering as nanodomains were observed on the surface of CD4 + CD25 high  regulatory T cells, these CD25 nanodomains were not co-localized with CD4 nanodomains. This result presented that the formation of these CD25 nanodomains on the surface of CD4 + CD25 high  T cells were not associated with the response of T cell receptor (TCR)/CD3-dependent signal transduction. In contrast, on the surface of CD4 + CD25 low  T cells, CD25 molecules distributed randomly without forming nanodomains while CD4 clustering as nanodomains can be observed; on the surface of CD8 + CD25 +  T cells, CD25 clustering as nanodomains and co-localization with CD8 nanodomains were observed. Collectively, above these results exhibited that TCR/CD3-based microdomains were indeed required for TCR/CD3-mediated T cells activation and enhanced the immune activity of CD4 + CD25 low  T cells or CD8 + CD25 +  T cells. In particular, it was found that the formation of CD25 nanodomains and their segregation from TCR/CD3 microdomains were the intrinsic capability of CD4 + CD25 high  T cells, suggesting this specific imaging feature of CD25 should be greatly associated with the regulatory activity of CD4 + CD25 high  T cells. Importantly, this novel NSOM/QD-based dual-color imaging system will provide a useful tool for the research of distribution-function relationship of cell-surface molecules.

Nanoscale Relationship Between CD4 and CD25 of T Cells Visualized with NSOM/QD-Based Dual-Color Imaging System