Fig. 2: Design and immunogenicity of a PfCSP nanoparticle immunogen.

a Schematic representation of immunogen 126, comprising the PfCSP epitope motifs KQPADG (brown), NPDP (green), NANP (blue) and NVDP (yellow) genetically fused to H. pylori apoferritin (gray) and PADRE (pink), separated by short linkers (black). Models of immunogen 126 displayed as both a monomer and as an assembled nanoparticle. The PfCSP epitope is presented externally on the nanoparticle surface. For better illustration, six monomers have been removed from the assembled nanoparticle, thus slicing through the immunogen, to reveal the PADRE epitopes within its core. b Size exclusion chromatogram and SDS-PAGE analysis of immunogen 126 (uncropped SDS-PAGE in Supplementary Fig. 3a). c Negative stain electron microscopy of immunogen 126. Scale bar—50 nm. d C57BL/6J mice were immunized with FL-PfCSP or immunogen 126 adjuvanted with SAS at day 0, 21 and 42. The serum IgG response against the PfCSP repeat (NANP) and junction (NPDP) was determined at the indicated timepoints. Immunization with SAS alone served as negative control. Pooled data of two independent experiments with 5 mice per group are shown. e Mice from the Kymouse™ platform were immunized with FL-PfCSP or immunogen 126 adjuvanted with SAS at day 0, 28 and 70. The serum IgG response against the PfCSP repeat (NANP) and the junction (NPDP) was measured at the indicated timepoints. Immunization with SAS alone served as control. One representative out of two independent experiments with 7 mice per group is shown. f IGHV3-33 (left) and paired IGHV3-33/IGKV1-5 (right) gene usage frequency among sorted GC CSP+ or GC CSP− cells isolated from lymph nodes of mice from the Kymouse™ platform 7 days after the third immunization with immunogen 126 adjuvanted with SAS. Data from naïve B cells are shown for comparison. FACS gating strategy is shown in Supplementary Fig. 6. Dots represent individual mice. Pooled data of two independent experiments are shown (CSP−: n = 6, CSP+: n = 4; naïve: n = 20). g Silent (S, gray) and replacement (red) mutations in VH3-33 antibodies of mice from the Kymouse™ platform. CDRs are marked in light gray. Arrows indicate positions H.31 and H.50 with strong selection for replacement mutations. h Capacity of pooled sera (diluted 1:400, collected 7 days after the last immunization) from mice of the Kymouse™ platform to inhibit the hepatocyte traversal activity of Pf sporozoites in vitro. Dots represent independent traversal assay experiments (n = 4). Arithmetic mean (d, e, h), median with length of the whiskers as multiple of IQR (f) and SEM (d, e) are indicated. Statistically significant differences were calculated by two-tailed Mann–Whitney test. In (d) and (e) statistical analyses were performed with data from day 50 and day 80, respectively (*P < 0.05; **P < 0.01; ***P < 0.001). Statistically non-significant differences are not indicated.