Fig. 5.

Activation of tyrosinase-specific T cells by tyrosinase-loaded monocyte-derived dendritic cells matured in the presence of different stimulatory cocktails. Monocytes from six donors were analyzed for the production of IL-12p70 after an initial 48 h culture with GM-CSF/IL-4 followed by harvest, wash, and thereafter 18 h with GM-CSF/IL-4 together with the combinations of IFNα, TNFγ, R848, GMP-grade poly I:C Hiltonol^®, different forms of LPS, and/or monophosphoryl Lipid A (MPLA). The mature monocyte-derived dendritic cells from the first three donors were then harvested, washed, and either kept unloaded or pulsed with tyrosinase or HCV peptide and co-cultured with allogeneic tyrosinase-specific TCR-transduced T cells. Specific activation was calculated by subtracting the IFNγ response against the control HCV peptide-loaded DC from the IFNγ response against the tyrosinase peptide-loaded DC (a). For the last three donors, tumor cell lysate from either tyrosinase-negative A375 cells or tyrosinase-transduced A375 was added during the maturation from immature to mature dendritic cells. Thereafter the dendritic cells were harvested, washed, and co-cultured with the tyrosinase-specific T cells. Specific activation was calculated by subtracting the IFNγ response against the control A375 lysate-loaded DC from the IFNγ response against the tyrosinase-expressing A375 lysate-loaded DC (b)