Figure 6

Induction of RCC-specific CTLs using RNA-transfected DCs as APCs. Monocyte-derived DCs were transfected with total RNA (5 μg/0.5 × 10⁶ DCs) isolated either from RCC-26 (A) or RCC-53 (B) tumor cell lines. After a maturation period of 48 h the DCs were co-cultivated with autologous PBMCs (PBMC/DC ratio = 8:1) for 10–12 days following three rounds of restimulation (PBMC/DC ratio = 4:1). In the case of RCC-53 specific CTLs (B) an additional round of restimulation was carried out using autologous LCLs transfected with RCC-53-derived total RNA. The PBMC/DC co-culture was supplemented with IL-7 and IL-2 as described in the Methods section. A) RCC-26-specific CTLs were harvested three days after the last round of restimulation and specific lysis was analyzed in a standard 4-h ⁵¹Cr-release assay. B) RCC-53-specific T cells were harvested 10 days after the last round of restimulation and specificity of the T cells was determined by measuring IFNγ in the 24 h supernatant media T cell/target cell co-cultures (1.5 × 10⁴ T cells : 1.5 × 10⁴ target cells per 200 μl).