Adeno-associated virus analysis by size exclusion chromatography within 3 minutes using short bio-inert columns made with 3 µm particles operated at high flowrates.

Adeno-associated virus (AAV) analytical characterization is crucial to the well-defined and reproducible production of human gene therapies utilizing the AAV vector modality. The establishment of analytical methods based upon technology platforms currently widely used by bio-therapeutic manufacturers, namely HPLC, will assist efforts to produce high quality AAV reproducibly and decrease chemical manufacturing and control challenges in method portability and reliability. AAV analysis by size exclusion chromatography (SEC) is currently practiced with columns and mobile phase conditions traditional to SEC of proteins. Here, an improved method to measure multiple AVV critical quality attributes (CQA) rapidly by SEC is explored. The use of short columns made with small particles at high flow rates resulted in up to 80 % reduction in analysis time and 66 % in sample consumption while maintaining reliable quantitation of AAV aggregate or high molecular weight (HMW) content. These results were demonstrated across four different AAV serotypes. Furthermore, critical AAV sample handling learnings are shared.
Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: James Y. Song has patent pending to Phenomenex. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
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