Extended Data Fig. 1 |. Production and characterization of HA-I53_dn5 components and nanoparticle immunogens.

a, SEC purification of seasonal HAs fused to I53_dn5B trimeric components, using a Superdex 200 Increase 10/300 GL column. b, Reducing and non-reducing SDS-PAGE of SEC-purified trimeric HA-I53_dn5B fusions, pentameric I53_dn5A component, and I53_dn5B trimer lacking fused HA. c, SEC purification of nanoparticle immunogens after in vitro assembly, including I53_dn5 lacking displayed antigen, using a Superose 6 Increase 10/300 GL column. The nanoparticle immunogens elute at the void volume of the column, while I53_dn5 is resolved. Residual, unassembled trimeric and pentameric components elute around 15 mL and 18 mL, respectively. d, Reducing and non-reducing SDS-PAGE of SEC-purified nanoparticle immunogens and I53_dn5. e, Antigenic characterization of purified nanoparticle immunogens by ELISA. Symbols indicate the specificity of each mAb. AUC, area under the curve. f, Analytical SEC of purified nanoparticle immunogens, compared to I53_dn5 nanoparticles lacking displayed antigen and trimeric H1-I53_dn5B, using a Sephacryl S-500 HR 16/60 column. g, Dynamic light scattering (DLS) of SEC-purified nanoparticle immunogens, including I53_dn5. D_h, hydrodynamic diameter; Pd, polydispersity. h, Representative electron micrograph of H1-I53_dn5 embedded in vitreous ice. Scale bar, 100 nm. i, 2D class averages obtained using single-particle cryo-EM. Scale bar, 20 nm. j, Gold-standard Fourier shell correlation curve for the H1-I53_dn5 density map presented in Fig. 1c. k, Gold-standard Fourier shell correlation curve for the localized reconstruction of H1 MI15 presented in Fig. 1c. All experiments except for electron microscopy data collection and processing were performed at least twice.