Journal of biological engineering · 2025
Baculovirus-mediated production and purification of ferritin nanoparticles for rift valley fever vaccine development.
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Rodrigues Margarida Q, Tambaclal Ashni, Kloss Brian, Alves Paula M, Roldão António
Аннотация
BACKGROUND: Rift Valley fever (RVF) is a WHO-prioritized zoonotic, vector-borne disease with no licensed prophylaxis available for humans, highlighting the need for effective vaccine strategies. Nanoparticle-based platforms for antigen presentation offer a promising approach for vaccine development.
RESULTS: In this work, we engineered ferritin (Ft) nanoparticles to display the immunogenic Gn domain of RVF virus (GnFt) and systematically assessed the production, purification, and physico-chemical properties of the purified nanoparticles. Baculovirus-based expression systems were evaluated in insect (Sf9, High-Five™, Tnao38, and Tnms42) and mammalian cells (HEK293 and CHO), revealing Sf9 cells as the most efficient host for producing GnFt nanoparticles. In addition, affinity-based chromatography was explored, yielding GnFt nanoparticles of > 95% purity (as assessed by SDS-PAGE) and an overall production yield of 0.2 mg/L culture. Biophysical characterization (e.g., high-performance liquid chromatography, dynamic light scattering, electron microscopy, and mass photometry) confirmed proper 24-mer nanoparticle assembly (1,344 kDa and 20 nm) and structural integrity. Binding affinity to Gn-targeting monoclonal antibodies was demonstrated by biolayer interferometry, with dissociation constants in the nM range, indicating retained antigenic functionality.
CONCLUSIONS: These findings demonstrate the successful development of a platform for producing structurally stable, pure, and functional Gn-presenting ferritin nanoparticles, supporting their potential use for RVF vaccine development.
RESULTS: In this work, we engineered ferritin (Ft) nanoparticles to display the immunogenic Gn domain of RVF virus (GnFt) and systematically assessed the production, purification, and physico-chemical properties of the purified nanoparticles. Baculovirus-based expression systems were evaluated in insect (Sf9, High-Five™, Tnao38, and Tnms42) and mammalian cells (HEK293 and CHO), revealing Sf9 cells as the most efficient host for producing GnFt nanoparticles. In addition, affinity-based chromatography was explored, yielding GnFt nanoparticles of > 95% purity (as assessed by SDS-PAGE) and an overall production yield of 0.2 mg/L culture. Biophysical characterization (e.g., high-performance liquid chromatography, dynamic light scattering, electron microscopy, and mass photometry) confirmed proper 24-mer nanoparticle assembly (1,344 kDa and 20 nm) and structural integrity. Binding affinity to Gn-targeting monoclonal antibodies was demonstrated by biolayer interferometry, with dissociation constants in the nM range, indicating retained antigenic functionality.
CONCLUSIONS: These findings demonstrate the successful development of a platform for producing structurally stable, pure, and functional Gn-presenting ferritin nanoparticles, supporting their potential use for RVF vaccine development.