In Vitro Assay to Sensitively Measure Tfr Suppressive Capacity and Tfh Stimulation of B Cell Responses

T follicular helper (Tfh) cells stimulate, whereas T follicular regulatory (Tfr) cells inhibit, B cell responses. Despite the potent immunoregulatory roles for Tfr cells in controlling the magnitude of antibody production, the precise mechanisms by which Tfr cells exert their suppressive effects are not yet clear. The lack of specific assays to assess Tfr cell function separately from differentiation has hindered progress in elucidating Tfr cell function. This is due, in part, to difficulty in separating Tfr cells from phenotypically similar, but functionally different, Tfh cells. Here we describe an in vitro approach for sensitively and quantitatively assessing the capacity of Tfr cells to suppress Tfh-mediated B cell antibody production utilizing both ELISA and flow cytometry to measure B cell responses. Beyond assessing Tfr function, this assay system can also be used to sensitively measure Tfh stimulatory capacity as well as B cell function.