Fig. 4. Immunisation with combinations of RH5, CyRPA and/or RIPR does not improve over immunisation with RH5 alone.

Wistar rats were immunised on days 0, 28 and 56. Terminal bleed was taken on day 70 (post-third dose). Doses of 2 µg RH5, and 20 µg CyRPA and RIPR antigen were used. For the “R + C+Ri” group, antigen doses were matched to the single antigen vaccination groups (2 µg RH5 + 20 µg CyRPA + 20 µg RIPR). For the “R + C+Ri Equimolar” group antigens were dose-matched to the RCR-complex (5.3 µg RH5 + 3.5 µg CyRPA + 11.1 µg). Day 70 serum IgG ELISA data (reported in µg/mL) shown against full-length (A) RH5 (red), (B) CyRPA (blue), and (C) RIPR (green). The dotted line corresponds to the median antigen-specific IgG response from the relevant group of single antigen-immunised animals. A summary of statistical analysis related to these panels can be found in Table S2. D Single-cycle GIA assays were performed using P. falciparum clone 3D7. Total IgG, purified from day 70 serum samples, was titrated in the GIA assay (see Supplementary Fig. 3B). GIA at 1 mg/mL total purified IgG was interpolated for each animal. The dotted line indicates the median RH5 GIA. Significance determined by one-way ANOVA with Dunnett’s multiple comparisons test versus RH5 group only, **** p < 0.0001. E Data from (D) were replotted against total antigen-specific IgG concentration in µg/mL as measured by ELISA in each purified total IgG sample (see Supplementary Fig. 3C). Each dataset was fitted with a Richard’s five-parameter dose-response curve with no constraints to ascertain the GIA assay EC₅₀. The dotted line shows the median result for the RH5 only group for comparison. Individual and median group responses (N = 6 per group) are shown in all panels. Significance was determined by one-way ANOVA of log-transformed data with Dunnett’s multiple comparisons test versus the RH5 group only. * p < 0.05, ** p < 0.001, *** p < 0.0001, **** p < 0.0001. Source data are provided as a Source Data file.