Precision of protein aggregation measurements by sedimentation velocity analytical ultracentrifugation in biopharmaceutical applications
Abstract
Sedimentation velocity analytical ultracentrifugation (SV-AUC) is routinely applied in biopharmaceutical development to measure levels of protein aggregation in protein products. SV-AUC is free from many limitations intrinsic to size exclusion chromatography (SEC) such as mobile phase and column interaction effects on protein self-association. Despite these clear advantages, SV-AUC exhibits lower precision measurements than corresponding measurements by SEC. The precision of SV-AUC is influenced by numerous factors, including sample characteristics, cell alignment, centerpiece quality, and data analysis approaches. In this study, we evaluate the precision of SV-AUC in its current practice utilizing a multilaboratory, multiproduct intermediate precision study. We then explore experimental approaches to improve SV-AUC measurement precision, with emphasis on utilization of high quality centerpieces.
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AUC is limited to purified samples without residual impurities or excipients and requires prior titration into the linear range 2 × 10^12-5 × 10^12 cp/mL.
“Sensitivity of the detectors and the risk of induced density gradients limit AUC to purified samples without residual impurities or excipients, and prior titration into the small linear range of the detector, 2 × 1012–5 × 1012 cp/mL, is needed.”
From Analytical methods for process and product characterization of recombinant adeno-associated virus-based gene therapies (2021) · cites this paper as [52]
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- Authors
- Gabrielson, J. P., Arthur, K. K., Stoner, M. R., Winn, B. C., Kendrick, B. S., Razinkov, V., Svitel, J., Jiang, Y., Voelker, P. J., Fernandes, C. A., & Ridgeway, R.
- Affiliations
- Amgen (United States) (US)
- Corresponding author
- Gabrielson, John P.
- ORCID
- Kendrick, Brent S.
- Venue
- Analytical Biochemistry
- Publisher
- Elsevier BV
- Published
- 2010-01
- Volume
- 396
- Issue
- 2
- Pages
- 231-241
- ISSN
- 0003-2697 (ISSN-L)
- OpenAlex
- W2047565601
- Cited by
- 56
- References
- 22
- Citations by year
- 2026: 1 · 2025: 5 · 2024: 5 · 2023: 3 · 2022: 1 · 2021: 2 · 2020: 3 · 2019: 4 · 2018: 3 · 2017: 3 · 2016: 4 · 2015: 7 · 2014: 3 · 2013: 3 · 2012: 4
- FWCI
- 1.20
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Analytical Biochemistry (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Amgen
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein purification and stability
- Life Sciences › Immunology and Microbiology › Immunology › Biosimilars and Bioanalytical Methods
- Physical Sciences › Chemistry › Analytical Chemistry › Analytical Methods in Pharmaceuticals
- Keywords
- protein aggregation, measurement precision, intermediate precision, cell alignment, biopharmaceutical products, protein self-association
- MeSH terms
- Chromatography, Gel, Protein Binding, Protein Conformation, Proteins, Ultracentrifugation
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