Rapid Determination of Full and Empty Adeno-Associated Virus Capsid Ratio by Capillary Isoelectric Focusing

Adeno-associated virus (AAV) is one of the most promising gene transfer vector types featuring long-term expression and low toxicity. The lack pathogenicity availability many serotypes augmented applicability AAV virions in therapy applications. recombinant capsid includes therapeutic protein-coding transgene as well a promoter to initiate translation poly A sequence portion for stabilization. Current manufacturing technologies, however, cannot guarantee generation only full capsids, i.e., including entire required genome. Partially filled empty capsids are also part product, decreasing this way efficacy safety upon clinical translation. Therefore, rapid, accurate QC friendly analysis ratio high importance during release testing. In paper, an automated capillary isoelectric focusing technique introduced, readily applicable biopharmaceutical industry fast efficient determination ratio. method reveals information about proportion partially capsids. For higher resolution (<0.1 pI unit), mixtures wide narrow range ampholytes were utilized. point peak area percentage reproducibility (RSD) mixed ampholyte assay 1.67% 2.45 %, respectively, requiring 65 nL sample volume per injection.