Efficient Selection of Recombinant Fluorescent Vaccinia Virus Strains and Rapid Virus Titer Determination by Using a Multi-Well Plate Imaging System
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cell Lines
2.2. Vaccinia Virus Strains
2.3. Plasmid
2.4. Recombinant Vaccinia Virus Generation by Homologous Recombination
2.5. Fluorescence-Based Viral Plaque Assay
2.6. Viral Plaque Assay for Statistical Analysis
2.7. Statistical Analysis
2.8. Software
3. Results
3.1. Generation, Selection, and Purification of the New L1c-Ig-Turbo-TK-eGFP Strain Expressing eGFP and Turbofp635 Proteins
3.2. Vaccinia Virus Titer Determination Based on Fluorescence-Dependent Plaque Assay
3.3. Development of a Plaque Assay for Non-Fluorescent rVACV on the Basis of Cell Lines Expressing Fluorescent Proteins
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Appendix A
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Plate No. | Total Number of Viral Plaques | Number of eGFP-Positive Plaques | Recombination Rate |
---|---|---|---|
1 | 522 | 3 | 0.57% |
2 | 550 | 5 | 0.91% |
3 | 567 | 3 | 0.53% |
4 | 493 | 4 | 0.81% |
5 | 487 | 6 | 1.23% |
6 | 475 | 3 | 0.63% |
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Ye, M.; Keicher, M.; Gentschev, I.; Szalay, A.A. Efficient Selection of Recombinant Fluorescent Vaccinia Virus Strains and Rapid Virus Titer Determination by Using a Multi-Well Plate Imaging System. Biomedicines 2021, 9, 1032. https://doi.org/10.3390/biomedicines9081032
Ye M, Keicher M, Gentschev I, Szalay AA. Efficient Selection of Recombinant Fluorescent Vaccinia Virus Strains and Rapid Virus Titer Determination by Using a Multi-Well Plate Imaging System. Biomedicines. 2021; 9(8):1032. https://doi.org/10.3390/biomedicines9081032
Chicago/Turabian StyleYe, Mingyu, Markus Keicher, Ivaylo Gentschev, and Aladar A. Szalay. 2021. "Efficient Selection of Recombinant Fluorescent Vaccinia Virus Strains and Rapid Virus Titer Determination by Using a Multi-Well Plate Imaging System" Biomedicines 9, no. 8: 1032. https://doi.org/10.3390/biomedicines9081032