Fig. 4

Priming and heterologous boosting of BALB/c mice using human–mouse chimeric anti-Clec9A Ab. a Schematic representation of the two new chimeric human–mouse anti-Clec9A-TpD-M2e constructs prepared from phage display Abs, compared with the rat anti-Clec9A-TpD-M2e construct. b Schematic representation of the experimental setup. BALB/c mice (n = 5 mice per group) were injected i.v. with 2 μg human–mouse chimeric anti-Clec9A-TpD-M2e (clones #2 or #19) or 2 μg rat anti-Clec9A-TpD-M2e (clone 10B4) in the presence of 5 nmol CpG. An unprimed control group was injected with 100 μl PBS. At day 28 after priming, each group of mice primed with the human–mouse chimeric Ab constructs were boosted with 2 μg rat anti-Clec9A-TpD-M2e (clone 10B4) and the groups of mice primed with the rat anti-Clec9A construct boosted with 2 μg human–mouse chimeric anti-Clec9A-TpD-M2e (clones #2 or 19) in the presence of 5 nmol CpG. Unprimed control groups were likewise injected at day 28 to give a primary response for direct comparison. c Serum samples were collected at the indicated time points post immunisation and the anti-M2e IgG response was measured by ELISA. Each point represents one individual mouse and the end point titre is shown as geometric mean with 95% CI. This experiment was performed twice and the results pooled (n = 10 mice per group). The boost responses were analysed by unpaired two-tailed Student's t-test and significance was indicated as ** p < 0.01, ***p < 0.001, ****p < 0.0001. The differences of Ab response between day 28 and at day 42 were indicated in Fig. 4c and the differences of Ab response between primed-boosted groups and unprimed-boosted groups at day 42 are followed; red open circle vs. black open triangle (**p = 0.0012), blue open circle vs. black open triangle (****p < 0.0001), red closed circle vs. red open triangle (***p = 0.0008), and blue closed circle vs. blue open triangle (**p = 0.0024)