ORN-stimulated DCs activate splenocytes to produce IFN-γ. BMDCs were stimulated with ORN 9.2dr PTO, PolyA PTO, or CpG ODN 1826 (A: 1 μg/mL; B: 0.1, 1, or 10 μg/mL) complexed with DOTAP for 6 hours, then washed and cocultured with naive splenocytes. (A) After an additional 18 hours of culture, the activation status of the splenic T-cell (CD3⁺ and CD4⁺) and B-cell (B220⁺, CD11c⁻) populations was determined by measuring expression of the early activation marker CD69. Representative data from one of 3 experiments are gated on CD3⁺, CD8⁺ T cells or B cells, respectively. Numbers indicate the percent of gated lymphocytes that are CD69⁺. (B) IFN-γ production by splenocytes after 18 hours of coculture was measured in the supernatant by ELISA. Data show the mean of triplicate samples ± SEM and are representative of 3 independent experiments. *P < .001; ns indicates not significant. The asterisk without brackets indicates comparison to unstimulated sample.