A simplified chromatography method for robust separation of full and empty AAV capsids: a case study with AAV2

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.