The purification and functional analysis of human CD4+CD25high regulatory T cells

Regulatory T cells were initially identified and isolated in the mouse, by virtue of their endogenous expression of CD25 (IL-2R alphachain) and shown to inhibit both the in vivo development of autoimmunity and the in vitro proliferation of nonregulatory, CD4+CD25- T cells. In contrast to mouse cells, human regulatory T cells are not purified by isolating all CD25-expressing CD4 T cells ex vivo. Such cells can be isolated by targeting only the small percentage of human CD4 T cells that express high levels of CD25. This is best achieved by FACS sorting using the level of CD25 expressed on CD4- T cells to place the gate for discriminating high expression of CD25. This unit provides two widely used methods to isolate (FACS) or to enrich (magnetic beads) human CD4+CD25+ regulatory T cells from blood, along with an in vitro coculture assay to measure the anergic and suppressive features of human CD4+CD25+ regulatory T cells.