Figure 6.

Dimeric CD3 aptamers enhance proliferation of suboptimally activated CD8+ OT-1 T cells

(A) Naive CD8 + OT-1 T cells were co-incubated with varying concentrations of SIINFEKL (high-affinity OT-1 peptide) and SIIGFEKL (low-affinity OT-1 peptide) and constant dose of CD3 aptamers. Rate of proliferation after 72 h was assayed via CFSE dilution. Proliferative effect of CD3 aptamers was only observed when there was suboptimal proliferation induced by the agonistic peptides. CD3 aptamers 1 and 12 enhanced the proliferative index of OT-1 T cells stimulated with both peptides, but with different patterns. Representative images of this assay are shown above. Independent experiments were repeated with similar results n > 3 times. (B) Naive OT-1 splenocytes were suboptimally activated with SIINFEKL (high-affinity OT-1 peptide) and SIIGFEKL (low-affinity OT-1 peptide), and CD3 aptamers and thymidine ³H incorporation was determined by scintillation. Data are represented as mean of n = 4 technical replicates of one experiment (mean ± SEM). One-way ANOVA followed by a Bonferroni post hoc test was performed. “SIIGFEKL + Apt Ctrl” vs. “SIIGFEKL + Apt 1” p value = 0.0002. “SIIGFEKL” vs. “SIIGFEKL + Apt 1” p value <0.0001. (C) Naive OT-1 splenocytes were suboptimally activated as in (B), and supernatant was collected to measured IFN-γ secretion by ELISA. Data are represented as mean of n = 4 technical replicates of one experiment (mean ± SEM). One-way ANOVA followed by a Bonferroni post hoc test was performed.