High resolution ES-DMA quantifies AAV capsid DNA content by electrical mobility to mass correlation

Recombinant Adeno-Associated Virus (rAAV) is the leading viral vector platform for gene therapy. A persistent challenge in rAAV manufacturing and quality control is accurate assessment of the DNA content of purified viral capsids. Empty particles, as well as 'partial' and over-filled species commonly contaminate even highly purified rAAV preparations. These impurities contribute to high production costs and reduced therapeutic efficacy, resulting in higher required doses and increased risks to patient health. Here we present high-resolution electrospray differential mobility analysis (ES-DMA) as a novel benchtop analytical modality for rAAV vector characterization. Sampling microliter volumes of analyte, this method achieves angstrom-scale particle sizing which we demonstrate is sufficient to resolve DNA encapsidation by rAAVs via differential electrical mobility. We benchmark this approach against charge detection mass spectrometry (CD-MS), an emerging gold standard for rAAV analytics, and identify a robust near-linear correlation between electrical mobility and mass of aerosolized rAAV particles. This relationship enables rapid identification and relative quantification of empty, full, and partial / over-filled rAAV capsids by ES-DMA.