Fig. 6. CD8− γδ T cells differentiation into CD8+ γδ T cells during in vitro culture.

MNCs isolated from spleen and IEL of Gn pigs (n = 3) infected with human norovirus GII.4 were sorted into three γδ T cell subsets by using MACS and FACS technologies. Each γδ T cell subset was cultured in vitro for 5 days with the stimulation of IPP plus IL-2 or mock stimulation. The culture supernatants were kept to measure IFN-γ, IL-10 and TGF-β by ELISA (See Fig. 5 legend) and the cells were collected for the Tcr1-N4 and CD8 staining. The left dot plot in panel (A) shows the purity of γδ T cells after enrichment by MACS and the right dot plot in panel (A) demonstrates the gating of three γδ T cell subsets to be collected by FACS. The table in panel (B) lists the mean frequencies of CD8 expression after 5 days in vitro culture of the CD2+CD8− and CD2−CD8− subsets from spleen and IEL.