Poly I:C–induced DC activation is inhibited by ssCpG-ODN. (A) Immature DCs were treated for 48 hours with the TLR ligands ssCpG-ODN, LPS, R848, or poly I:C. Flow cytometry was used to measure CD80 and CD86 expression. The results are depicted as mean fluorescent intensity (MFI; mean ± SEM, n = 6). Significant differences compared with medium control were assessed by repeated measures ANOVA with Bonferroni multiple comparison; **P < .01 and ***P < .001. NS indicates nonsignificant. (B) DCs were treated for 48 hours with TLR ligands LPS, R848, and poly I:C as single treatment or in combination with ssCpG-ODN. DC maturation was determined by assessing CD80 and CD86. Flow cytometry results are depicted as the MFI (mean ± SEM values, n = 7 for LPS and R848, and n = 13 for medium and poly I:C treatments). Significant differences comparing with or without ssCpG-ODN were assessed by noparametric Mann-Whitney 2-tailed test; ***P < .001. (C) The expression of CD1a and up-regulation of CD80 and CD86 was measured with flow cytometry on DCs 48 hours after ligand addition and displayed as a representative figure. (D) Dose-response measurement (MFI mean ± SEM values, n = 7) of the ssCpG-ODN inhibitory effect on poly I:C–induced DC maturation. (E) Secretion of proinflammatory cytokines IL-12p40, IL-12p70, and TNF-α in the supernatants at 24 hours was measured using Bio-Plex technology. Significant differences comparing with or without ssCpG-ODN (n = 8) were assessed by nonparametric Mann-Whitney 2-tailed test; *P < .05 and **P < .01.