Characterization and quantification of adeno-associated virus capsid-loading states by multi-wavelength analytical ultracentrifugation with UltraScan

Amy Henrickson, Xiaozhe Ding, Austin G. Seal, Zhe Qu, Lauren Tomlinson, John Forsey, Viviana Gradinaru, Kazuhiro Oka, Borries Demeler

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

Aim: We present multi-wavelength (MW) analytical ultracentrifugation (AUC) methods offering superior accuracy for adeno-associated virus characterization and quantification. Methods: Experimental design guidelines are presented for MW sedimentation velocity and analytical buoyant density equilibrium AUC. Results: Our results were compared with dual-wavelength AUC, transmission electron microscopy and mass photometry. In contrast to dual-wavelength AUC, MW-AUC correctly quantifies adeno-associated virus capsid ratios and identifies contaminants. In contrast to transmission electron microscopy, partially filled capsids can also be detected and quantified. In contrast to mass photometry, first-principle results are obtained. Conclusion: Our study demonstrates the improved information provided by MW-AUC, highlighting the utility of several recently integrated UltraScan programs, and reinforces AUC as the gold-standard analysis for viral vectors.

Original languageEnglish
Pages (from-to)1519-1534
Number of pages16
JournalNanomedicine (London, England)
Volume18
Issue number22
DOIs
StatePublished - Sep 1 2023

Keywords

  • UltraScan
  • adeno-associated virus
  • analytical buoyant density gradient equilibrium
  • gene therapy
  • multi-wavelength analytical ultracentrifugation
  • sedimentation velocity
  • viral capsid quantification
  • viral vector

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