Immunostimulatory ORNs induce antigen-specific cytotoxicity in vivo. Mice were immunized as described in Figure 5 with 50 μg OVA together with ORN 9.2dr PTO, PolyA PTO, or CpG ODN twice at a 14-day interval. One week after the second immunization, spleen cells were isolated. Cytotoxic potential was demonstrated in vivo by specific lysis of labeled splenocytes loaded with OVA_257-264 peptide compared to unloaded splenocytes. Numbers above boxed areas indicate the percent of CFSE⁺ cells that are loaded with peptide (CFSE^high, right box) or unloaded (CFSE^low, left box). (A) Representative data from one of 2 experiments are gated on CFSE⁺ cells. (B) Data show the mean values of individual mice (OVA, OVA + ORN: n = 5; no OVA, OVA + CpG: n = 3) for one experiment ± SEM. Results are representative of 2 independent experiments. *P < .01; **P < .001; ns indicates not significant. The asterisk without brackets indicates comparison to OVA group.