Preclinical Establishment of a Divalent Vaccine against SARS-CoV-2
Abstract
:1. Introduction
2. Materials and Methods
2.1. Study Rationale
2.2. Animals, Blood Sampling and Tissue Processing
2.3. Construction and Heterologous Expression of the Fusion Protein
2.4. Directional Cloning into the Gateway pEntry/D-TOPO Vector
2.5. Insertion into Baculovirus and Amplification in Insect Cells
2.6. ‘VieVac’ Purification from Insect Cells
2.7. Evaluation of Protein Expression by Immunofluorescence
2.8. Protein Production and Purification from E. coli
2.9. Immunoblotting of ‘VieVac’ Protein with Immune Sera
2.10. Protein Adsorption to Imject™ Alum Adjuvant
2.11. Mouse Immunization
2.12. ELISA
2.13. RNA Isolation from Blood and Spleen
2.14. Quantitative PCR
2.15. Ex Vivo T-Cell Stimulation with N- and S-Specific Peptides and CD44 Evaluation by Cell Surface Flow Cytometry
2.16. Immunohistochemistry
2.17. Statistical Analysis
3. Results
3.1. Protein Production, Characterization and Purification
3.2. Imject™ Alum/’VieVac’ Induces IgGs to Both Nucleocapsid and RBD
3.3. AddaVax™ with ‘VieVac’ Primes IgG Production to Both Nucleocapsid and RBD
3.4. Insect-Cell-Produced AddaVax™/VieVac Generates Effector Cell Immunoreactivity
3.5. CD4+ and CD8+ T-Cell Responses Evaluated by CD44 Cell Surface Upregulation in Ex Vivo-Stimulated Spleen Cells of AddaVax™-/VieVac-Challenged Mice
3.6. Lack of Adverse Neuropathology in Addavax™-/VieVac-Injected Mice
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Hevesi, Z.; Gerges, D.A.; Kapps, S.; Freire, R.; Schmidt, S.; Pollak, D.D.; Schmetterer, K.; Frey, T.; Lang, R.; Winnicki, W.; et al. Preclinical Establishment of a Divalent Vaccine against SARS-CoV-2. Vaccines 2022, 10, 516. https://doi.org/10.3390/vaccines10040516
Hevesi Z, Gerges DA, Kapps S, Freire R, Schmidt S, Pollak DD, Schmetterer K, Frey T, Lang R, Winnicki W, et al. Preclinical Establishment of a Divalent Vaccine against SARS-CoV-2. Vaccines. 2022; 10(4):516. https://doi.org/10.3390/vaccines10040516
Chicago/Turabian StyleHevesi, Zsofia, Daniela Anna Gerges, Sebastian Kapps, Raimundo Freire, Sophie Schmidt, Daniela D. Pollak, Klaus Schmetterer, Tobias Frey, Rita Lang, Wolfgang Winnicki, and et al. 2022. "Preclinical Establishment of a Divalent Vaccine against SARS-CoV-2" Vaccines 10, no. 4: 516. https://doi.org/10.3390/vaccines10040516
APA StyleHevesi, Z., Gerges, D. A., Kapps, S., Freire, R., Schmidt, S., Pollak, D. D., Schmetterer, K., Frey, T., Lang, R., Winnicki, W., Schmidt, A., Harkany, T., & Wagner, L. (2022). Preclinical Establishment of a Divalent Vaccine against SARS-CoV-2. Vaccines, 10(4), 516. https://doi.org/10.3390/vaccines10040516