Identification of Foxp3⁺ T follicular regulatory (Tfr) cells by flow cytometry

Flow cytometry is a technology that allows multiparametric analysis of individual cells. As a consequence, it is among the most commonly used tools for the study of immune cells. It is useful both for the study of ex vivo cell populations isolated from experimental animals or human tissue and for characterizing the phenotype of cultured cells. The phenotypic analysis is based on antibodies associated to different fluorophores that specifically bind to key molecules. Genetically modified mouse strains that express a reporter gene under the control of a promoter of interest offer an important alternative for the staining of intracellular molecules without the need to permeabilize the cell membrane. In this chapter, we describe how Foxp3(+) follicular regulatory T (Tfr) cells, a population of regulatory T (Treg) cells related to T follicular helper (Tfh) cells and involved in the regulation of germinal centers (GC), can be identified by flow cytometry.