A simplified chromatography method for robust separation of full and empty AAV capsids: a case study with AAV2
Cell & Gene Therapy Insights 2025; 11(9), 743–0
DOI: 10.18609/cgti.2025.083
The inefficiency of AAV to package therapeutic genes results in the production of numerous empty or partially filled capsids, affecting the efficacy and safety of AAV-based therapies. Developing a scalable purification method to separate full and empty AAV particles remains challenging due to their similar properties.
In this study, a streamlined AAV2 downstream was developed with particular focus on the polish step. Affinity capture resulted in 95% recovery of AAV2-GFP functional particles. Subsequently, a conductivity-based strategy with an isocratic step hold with Thermo Scientific™ POROS™ XQ strong anion exchange resin was used to increase separation between empty and full capsids. Full capsids were enriched 5-fold and reduced empty capsids 4-fold. This impurity reduction minimizes aggregation risks and immune-mediated toxicities. This approach with a strong anion exchange resin offers an easily developable, scalable, and robust solution for AAV manufacturing, essential for supporting the production of safe and effective gene therapies.
POROS: Pharmaceutical Grade Reagent. For Manufacturing and Laboratory Use Only. CaptureSelect: For Research Use or Further Manufacturing. Not for diagnostic use or direct administration in humans or animals.