Figure 2.

Pig antigen-responding T cells can be expanded and visualized after monocyte-derived dendritic cell (MoDC) restimulation. (A) Experimental setup. Purified blood CD14⁺ monocytes were differentiated into MoDC for 7 days in the presence of GM-CSF and interleukin-4. MoDCs were primed with 40 µg/ml Candida albicans lysate and maturation was triggered by TNF-α and LPS treatment for 1 day. In parallel, PBMCs were CFSE-labeled and expanded in the presence of 20 µg/ml Candida antigen for 7 days followed by a 2-day resting phase. Expanded lymphocytes were restimulated with autologous, primed MoDC (MoDC:T cell ratio, 1:5) and CD154 expression was assessed according to the gating strategy in (B). CD154 expression of CD4⁺CFSE^low (proliferated cells, lower left plot) was compared to unproliferated CD4⁺CFSE^high (lower middle, red plot) and proliferated CD4⁻CD8α⁺CFSE^low T cells (lower right, blue plot). (C) As control, CD154⁺ frequency of Cand expanded PBMC without DC restimulation (w/o DC ctrl.) is shown exemplarily [gated on CD4⁺CFSE^low as depicted in panel (B)]. (D) CD154⁺ T cell frequencies after restimulating expanded T cells with unprimed (w/o) or Candida antigen-primed (Cand) MoDC (n = 6, Mann–Whitney test, **p = 0.0043).