Characterization of anti-PCV2 specific T lymphocytes. (A) Anti-CD4 and -CD8 mAbs reduce IFN-γ SC. PBMC from PCV2-immune animals were treated with mAbs against the CD4 and CD8 T cell receptors, or anti-CD1 as control for 1 h prior to PCV2 or mock antigen re-stimulation for 5 days (as in Fig. 5). The IFN-γ SC were measured by ELISPOT assay, and calculated per 106 cells. Mean values of triplicates of one representative experiment +/- SD are shown. (B and C) IFN-γ SC detected by flow cytometry. PBMC from PCV2-infected piglets (3 months after infection) were re-stimulated with PCV2 or mock antigen for 5 days prior to staining for the presence of intracellular IFN-γ. The cells were also labelled for the surface markers CD8 to differentiate MHC class I restricted CTLs as CD8hi cells from Th cells located within the CD8-/lo cell population. The latter includes also NK cells and γ/δ T cells. Values shown on the y-axis represent the percentage of IFN-γ positive cells. The symbols represent the individual animals. In C the percentage of the IFN-γ positive cells within the blasting cells defined by gating on cells with high forward scatter is shown. For B and C, monocytic cells were excluded based on their CD172a expression.