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Related Experiment Video

Updated: Sep 12, 2025

Author Spotlight: Advancing Gene Therapy Research with High-Titer Adeno-Associated Virus Vector Production
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Orthogonal approaches to AAV vector characterization: Validating quantitative TEM for partially filled particles.

Mattin Campos1, Dorleta Otaegui1, Taeyang Jung2

  • 1Viralgen SL, Paseo Mikeletegi, 8, 20009 San Sebastián, Spain.

Molecular Therapy. Methods & Clinical Development
|August 8, 2025
PubMed
Summary

Quantitative electron microscopy (QuTEM) accurately distinguishes full, partial, and empty adeno-associated virus (AAV) capsids. This method offers a reliable approach for AAV characterization, crucial for effective gene therapy development.

Keywords:
AAVMPSEC-HPLCSV-AUCTEMadeno-associated virusmass photometryscAAVsedimentation velocity analytical ultracentrifugationself-complementarysingle-stranded AAVsize exclusion chromatographyssAAVtransmission electron microscopy

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Area of Science:

  • Gene Therapy
  • Virology
  • Biotechnology

Background:

  • Adeno-associated viruses (AAVs) are key vectors in gene therapy.
  • Empty or partially filled capsids in AAV preparations reduce therapeutic efficacy.
  • Accurate characterization of AAV vector content is essential for quality control.

Purpose of the Study:

  • To present and validate quantitative electron microscopy (QuTEM) for AAV capsid characterization.
  • To compare QuTEM with analytical ultracentrifugation (AUC), mass photometry (MP), and SEC-HPLC.
  • To assess the effectiveness of these methods in evaluating AAV vector content.

Main Methods:

  • Quantitative electron microscopy (QuTEM) to distinguish AAV capsids by internal density.
  • Comparison of QuTEM with AUC, MP, and SEC-HPLC.
  • Production and evaluation of self-complementary (scAAV) and single-stranded (ssAAV) AAV vectors.

Main Results:

  • QuTEM reliably quantified AAV populations with high concordance to MP and AUC data.
  • QuTEM provided superior granularity by visualizing native viral capsids.
  • The study evaluated encapsidation efficiency for scAAV and ssAAV vectors.

Conclusions:

  • QuTEM is a robust and potentially gold-standard method for AAV characterization.
  • Integrated analytical approaches are vital for optimizing AAV production and ensuring gene therapy consistency.
  • Accurate AAV capsid quantification is critical for therapeutic efficacy.