A modular approach to suppression assays: TLR ligands, conditioned medium, and viral infection

The suppressive function of regulatory T-cells (Treg) requires precise control to allow an efficient adaption of the T-cell response to the requirements of the immune defense. In the setting of infection, an abrogation of the suppressive effect of Treg on the activation and proliferation of T-effector (Teff) cells is a central precondition to allow fast and efficient clearance of the infectious agent. Experimentally, the suppressive function of Treg on Teff can be indirectly measured in coculture proliferation assays. This versatile tool provides a readout of T cell proliferation in the presence of Treg through the measurement of a proliferation marker such as the incorporation of radioactively labeled thymidine ((3)H Thymidine), carboxyfluorescein succinimidyl ester (CFSE) or 5-Bromo-2'-deoxyuridine (BrdU). In a modular approach, the culture conditions can thereby be adapted to evaluate the effect of any cell type, live and inactivated microorganisms, molecularly defined immunostimulatory ligands, and cytokines on the interplay of Teff and Treg function. Here, we demonstrate how the suppression assay can be used as a multifunctional tool to provide insights into the interaction of Treg with Teff under a variety of conditions in vitro.