Detection of protein aggregates by sedimentation velocity analytical ultracentrifugation (SV-AUC): Sources of variability and their relative importance

✍ Scribed by Kelly K. Arthur; John P. Gabrielson; Brent S. Kendrick; Michael R. Stoner

• Publisher: John Wiley and Sons
• Year: 2009
• Tongue: English
• Weight: 305 KB
• Volume: 98
• Category: Article
• ISSN: 0022-3549
• DOI: 10.1002/jps.21654

✦ Synopsis

Sedimentation velocity analytical ultracentrifugation (SV-AUC) has found application in the biopharmaceutical industry as a method of detecting and quantifying protein aggregates. While the technique offers several advantages (i.e., matrix-free separation and minimal sample handling), its results exhibit a high degree of variability relative to orthogonal size-sensitive separation techniques such as size exclusion chromatography (SEC). The goal of this work is to characterize and quantify the sources of variability that affect SV-AUC results, particularly size distributions for a monoclonal antibody monomer/dimer system. Contributions of individual factors to the overall variability are examined. Results demonstrate that alignment of sample cells to the center of rotation is the most significant contributing factor to overall variability. The relative importance of other factors (e.g., temperature equilibration, time-invariant noise, meniscus misplacement, etc.) are quantified and discussed.