A Pentavalent Epstein-Barr Virus-Like Particle Vaccine Elicits High Titers of Neutralizing Antibodies against Epstein-Barr Virus Infection in Immunized Rabbits
Abstract
:1. Introduction
2. Methods
2.1. Animals and Ethics Statement
2.2. Cell Lines
2.3. Antibodies
2.4. Virus Production and Purification
2.5. Plasmids
2.6. Transfection of gp350-F-gB-F-gp42-gL-gH-F and Generation of Stable CHO Cell Line
2.7. Transfection, Generation, and Purification of EBV-LPs
2.8. SDS-PAGE, Coomassie Staining, and Immunoblotting
2.9. Transmission Electron Microcopy (TEM)
2.10. Recombinant EBV Proteins
2.11. Immunization of Rabbits
2.12. Determination of Glycoprotein-Specific Antibody Titers in Serum of Immunized Rabbits
2.13. Purification of IgG Antibodies from Rabbit Sera, and Determination of Glycoprotein-Specific Antibody Titers
2.14. EBV-eGFP Neutralization Assay in B cells and Epithelial Cells
2.15. Statistical Analysis
3. Results
3.1. Construction, Purification, and Characterization of EBV-LP Vaccine Candidate That Incorporates Five EBV Glycoproteins
3.2. Pentavalent EBV-LPs Are Immunogenic and Elicit EBV-Glycoprotein-Specific Antibodies in Rabbits
3.3. EBV-LPs Produce Robust nAb Responses in Immunized Rabbits That Prevent EBV Infection of Both B Cell and Epithelial Cell Lines
4. Discussion
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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2A Position | Nucleotide Sequence | Amino Acid Sequence |
---|---|---|
gp350-gB | GCTACTAACTTCAGCCTGCTGAAGCAGGCTGGAGACGTGGAGGAGAACCCTGGACC | A T N F S L L K Q A G D V E E N P G P |
gB-g42 | GCCACCAATTTCTCGTTACTTAAACAAGCGGGTGACGTTGAAGAGAATCCGGGACCT | A T N F S L L K Q A G D V E E N P G P |
g42-gL | GCGACTAACTTCTCATTGTTGAAACAGGCAGGAGATGTCGAAGAGAACCCTGGTCCA | A T N F S L L K Q A G D V E E N P G P |
gL-gH | GCAACGAATTTCTCCCTTCTAAAGCAAGCCGGTGACGTGGAGGAGAATCCCGGACCC | A T N F S L L K Q A G D V E E N P G P |
Primer Name | Primer Sequence (5’-3’) |
---|---|
Cloning Primers | |
EBV gH-gL-Fc-His FWD | AAAAAGCGGCCGCGCCACCATGCGTGCTGTTGGTGTATTTC |
EBV gH-gL-Fc-His REV | AAAAAACTAGTGTGTGCTCTTTCTTCATACAGG |
Sequencing primers | |
FC-Hisseqprimer4 | GCTTTAATAAGATCTCTAG |
FC-Hisseqprimer5 | TGCTGGGCACGGTGGGCATG |
FC-Hisseqprimer6 | GGGTCTTTTCTGCAGAAGCTTG |
IC50 *(µg/mL) | |||||
---|---|---|---|---|---|
Cell-Line | gp350 | UV-EBV | EBV-LPs | mAb 72A1 | mAb 2L10 |
HEK-293 | 5.67 | 3.11 | 2.85 | 6.25 | nd |
Raji | 8.97 | 3.42 | 3.71 | 4.81 | nd |
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Escalante, G.M.; Foley, J.; Mutsvunguma, L.Z.; Rodriguez, E.; Mulama, D.H.; Muniraju, M.; Ye, P.; Barasa, A.K.; Ogembo, J.G. A Pentavalent Epstein-Barr Virus-Like Particle Vaccine Elicits High Titers of Neutralizing Antibodies against Epstein-Barr Virus Infection in Immunized Rabbits. Vaccines 2020, 8, 169. https://doi.org/10.3390/vaccines8020169
Escalante GM, Foley J, Mutsvunguma LZ, Rodriguez E, Mulama DH, Muniraju M, Ye P, Barasa AK, Ogembo JG. A Pentavalent Epstein-Barr Virus-Like Particle Vaccine Elicits High Titers of Neutralizing Antibodies against Epstein-Barr Virus Infection in Immunized Rabbits. Vaccines. 2020; 8(2):169. https://doi.org/10.3390/vaccines8020169
Chicago/Turabian StyleEscalante, Gabriela M., Joslyn Foley, Lorraine Z. Mutsvunguma, Esther Rodriguez, David H. Mulama, Murali Muniraju, Peng Ye, Anne K. Barasa, and Javier Gordon Ogembo. 2020. "A Pentavalent Epstein-Barr Virus-Like Particle Vaccine Elicits High Titers of Neutralizing Antibodies against Epstein-Barr Virus Infection in Immunized Rabbits" Vaccines 8, no. 2: 169. https://doi.org/10.3390/vaccines8020169
APA StyleEscalante, G. M., Foley, J., Mutsvunguma, L. Z., Rodriguez, E., Mulama, D. H., Muniraju, M., Ye, P., Barasa, A. K., & Ogembo, J. G. (2020). A Pentavalent Epstein-Barr Virus-Like Particle Vaccine Elicits High Titers of Neutralizing Antibodies against Epstein-Barr Virus Infection in Immunized Rabbits. Vaccines, 8(2), 169. https://doi.org/10.3390/vaccines8020169