Continuous Affinity Purification of Adeno-Associated Virus Using Periodic Counter-Current Chromatography
Abstract
:1. Introduction
2. Materials and Methods
2.1. AAV Production
2.2. Screening of Elution Buffer
2.3. Batch Chromatography and Frontal Experiments
2.4. 4C-PCC Experiments
2.5. Analytical Quantification Methods
2.5.1. Total Particle Quantification
2.5.2. Genome Copies Quantification
2.5.3. Total Protein and ds-DNA
2.5.4. SDS-PAGE and Western Blot
2.5.5. Transmission Electron Microscopy
2.6. Chromatography Model
3. Results
3.1. Optimization of Elution Buffer
3.2. Breakthrough Curves
3.3. Chromatography Model of 4C-PCC Loading Zone
3.4. 4C-PCC Affinity Capture Process
4. Discussion
4.1. High Elution Recovery Yield Obtained after Buffer Screening
4.2. Modelling of the Breakthrough Curves Improves PCC Process Development
4.3. Continuous Affinity Capture Using 4C-PCC
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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Mendes, J.P.; Bergman, M.; Solbrand, A.; Peixoto, C.; Carrondo, M.J.T.; Silva, R.J.S. Continuous Affinity Purification of Adeno-Associated Virus Using Periodic Counter-Current Chromatography. Pharmaceutics 2022, 14, 1346. https://doi.org/10.3390/pharmaceutics14071346
Mendes JP, Bergman M, Solbrand A, Peixoto C, Carrondo MJT, Silva RJS. Continuous Affinity Purification of Adeno-Associated Virus Using Periodic Counter-Current Chromatography. Pharmaceutics. 2022; 14(7):1346. https://doi.org/10.3390/pharmaceutics14071346
Chicago/Turabian StyleMendes, João P., Magnus Bergman, Anita Solbrand, Cristina Peixoto, Manuel J. T. Carrondo, and Ricardo J. S. Silva. 2022. "Continuous Affinity Purification of Adeno-Associated Virus Using Periodic Counter-Current Chromatography" Pharmaceutics 14, no. 7: 1346. https://doi.org/10.3390/pharmaceutics14071346
APA StyleMendes, J. P., Bergman, M., Solbrand, A., Peixoto, C., Carrondo, M. J. T., & Silva, R. J. S. (2022). Continuous Affinity Purification of Adeno-Associated Virus Using Periodic Counter-Current Chromatography. Pharmaceutics, 14(7), 1346. https://doi.org/10.3390/pharmaceutics14071346