Fig. 1: Design and construction of ICO-RBD nanovaccines.

a Schematic illustration of the “engraving-printing” strategy to decorate RBD antigens onto ICO. The genomic DNA of M13mp18 bacteriophage was annealed and folded together with staple and capture strands (with polyA overhangs) to form the ICO nanocore. The SpyTag-N3 and DBCO-polyT were covalently linked through click chemistry to form the SpyTag-polyT conjugates. Next, the SpyTag peptides were engraved onto ICO through hybridization between polyT and polyA. The fusion proteins, SpyCatcher-RBD (prototype), were printed onto ICO through the spontaneous covalent isopeptide bond formation between SpyCatcher and SpyTag. b Schematic illustration of the optimization screening of surface antigen patterns based on the efficiency of the nanovaccines to activate RBD-specific IgG-BCR in vitro and the evaluation of efficient short- and long-term immune responses in vivo. Various surface antigen patterns with different parameters, including antigen spacing (10–40 nm), antigen copies within clusters (1–5) and cluster numbers (1–12), were screened. Characterization of ICO, including TEM (c), AFM (d) and DLS (n = 3 independent experiments) (e) analysis. Scale bar (white), 1 μm. Scale bar (black), 100 nm. f Gel electrophoresis of genomic DNA of M13mp18 bacteriophage and ICO after incubation in 20% FBS for different time. Gel electrophoresis (g) and loading efficiency of ICO-RBD nanovaccines surface-modified with 12, 24, 36, 48 or 60 RBD antigens (n = 4 independent experiments) (h). These experiments (c, d, f, g) were repeated three times independently with similar results. The data were processed on GraphPad Prism 8 and are presented as the mean ± SD. Source data are provided as a Source Data file.