Polarizing Cytokines for Human Th9 Cell Differentiation.

CD4+ T helper (Th) cell subset generation in vivo requires T cell receptor activation and surface CD28 co-stimulation in the presence of one or more cytokines. Similarly, Th cells can be generated in vitro by activating naïve CD4+CD25- T cells with plate bound-anti-CD3 monoclonal antibody (mAb) (pbCD3) and soluble-anti-CD28 mAb (sCD28) in the presence of polarizing recombinant (r) cytokines and anti-cytokine mAbs. In comparison to in vitro CD4+CD25- T cells, memory CD4+CD25-CD45RO+ T cells have been shown to convert to Th9 cells more efficiently. Here, protocol for in vitro generation of human Th9 cells by activating CD4+CD25-CD45RO+ memory T cells with pbCD3 and sCD28 in the presence of polarizing recombinant interleukin-4 (rIL-4) and transforming growth factor (rTGF-β) is described.