Fig. 3
HeV-GS586N elicits strong humoral immune responses against NiV and HeV in both mice and nonhuman primates. a Female BALB/c mice (6–8 weeks old, n = 6 per group) were immunized with 10 μg of NiV-G, HeV-G, or HeV-GS586N protein supplemented with 200 μg of aluminum hydroxide and 20 μg of CpG 1826 on days 0 and 21. Mice immunized with PBS served as negative controls. On day 42 after the prime immunization, serum neutralizing antibodies were measured via a Luminex-based competitive inhibition assay against NiV-Bangladesh (Bd NiV), NiV-Malaysia (My NiV), HeV, and HeV2. b Male cynomolgus monkeys (4–5 years old, n = 2 per group) were immunized with 50 μg of HeV-G or HeV-GS586N protein formulated with 1 mg of aluminum hydroxide and 100 μg of CpG 2006 on days 0 and 21. Serum binding antibody responses were assessed via ELISA (c, d, two technical replicates). Neutralizing activity was measured via pseudovirus neutralization assays (e, f, three technical replicates). Authentic virus neutralization assays were performed in a BSL-4 laboratory (g, h, four technical replicates). Receptor-binding inhibition was measured via Luminex-based competitive inhibition assays (i, two technical replicates). The IC50 values represent the dilution at which 50% infection or receptor binding is inhibited. NT50 values represent the reciprocal serum dilution at which 50% of virus infection is inhibited. The error bars indicate the standard deviation (SD) of biological replicates. Statistical significance was determined via unpaired two-tailed Student’s t tests, with *P < 0.05; **P < 0.01; and ***P < 0.001
