ArticleMolecular therapy. Methods & clinical development2020
A qPCR Method for AAV Genome Titer with ddPCR-Level of Accuracy and Precision.
Article in Molecular therapy. Methods & clinical development, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 25 papers, 1 of them a synthesis that pooled it.
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Who cites it
25 citing papers in PubMed, 1 synthesis or guideline pooled it.
- What's new and what's next for gene therapy in Pompe disease?Expert opinion on biological therapy · 2022Pooled it
- Chronologically distributed transfection improves AAV2 and AAV2/8 capsid filling and reveals assembly schedule divergence.Molecular therapy. Methods & clinical development · 2025Article
- Development of an HEK293 Suspension Cell Culture Medium, Transient Transfection Optimization Workflow, and Analytics for Batch rAAV Manufacturing.Biotechnology and bioengineering · 2025Article
- Impact of DNase digestion on titer measurements of engineered adeno-associated virus serotypes.Molecular therapy. Methods & clinical development · 2025Article
- Tracking Particle-Encapsulated DNA Across the Anion-Exchange Chromatography Fractions of Recombinant Adeno-Associated Virus Using Droplet Digital PCR and High-Throughput Sequencing.Biotechnology journal · 2025Article
- Quality assessment strategy development and analytical method selection of GMP grade biological drugs for gene and cell therapy.BBA advances · 2025Review
- Implementing a robust platform analytical procedure for measuring adeno-associated virus vector genome titer.Molecular therapy. Methods & clinical development · 2024Article
- An amplification-free CRISPR-Cas12a assay for titer determination and composition analysis of the rAAV genome.Molecular therapy. Methods & clinical development · 2024Article
- A Novel Method for Separating Full and Empty Adeno-Associated Viral Capsids Using Ultrafiltration.Membranes · 2024Article
- An autonucleolytic suspension HEK293F host cell line for high-titer serum-free AAV5 and AAV9 production with reduced levels of DNA impurity.Molecular therapy. Methods & clinical development · 2024Article
- Characterization of AAV vectors: A review of analytical techniques and critical quality attributes.Molecular therapy. Methods & clinical development · 2024Review
- In-Depth Comparison of Adeno-Associated Virus Containing Fractions after CsCl Ultracentrifugation Gradient Separation.Viruses · 2024Article
- Assessment of Key Factors Impacting Variability in AAV Vector Genome Titration by Digital PCR.International journal of molecular sciences · 2024Article
- Adeno-associated virus genome quantification with amplification-free CRISPR-Cas12a.Gene therapy · 2024Article
- Distributional comparison of different AAV vectors after unilateral cochlear administration.Gene therapy · 2024Article
- Concise Analysis of Single-Stranded DNA of Recombinant Adeno-Associated Virus By Automated Electrophoresis System.Human gene therapy · 2024Article
- PCR-based analytics of gene therapies using adeno-associated virus vectors: Considerations for cGMP method development.Molecular therapy. Methods & clinical development · 2023Review
- Design space determination to optimize DNA complexation and full capsid formation in transient rAAV manufacturing.Biotechnology and bioengineering · 2023Article
- Reversed Phase-Liquid Chromatography for Recombinant AAV Genome Integrity Assessment.Analytical chemistry · 2023Article
- Insight and Development of Advanced Recombinant Adeno-Associated Virus Analysis Tools Exploiting Single-Particle Quantification by Multidimensional Droplet Digital PCR.Human gene therapy · 2022Article
Corrections and comments
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Authors and funding
5 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Recombinant adeno-associated virus (rAAV) is one of the main vectors used in gene therapy. An accurate genome titer is not only critical for clinical dosing, but also a prerequisite for many analytical assays for AAV product characterization. AAV genome titer is traditionally determined by qPCR; however, assay precision is not optimal despite extensive efforts. More recently, droplet digital PCR (ddPCR) emerged as a powerful alternative that offers excellent accuracy and precision. However, currently ddPCR is not as widely available as qPCR and operates at a lower throughput and a higher cost. In this paper, we introduce an improved qPCR method with two major optimizations: (1) using an AAV reference material as qPCR standard instead of plasmid DNA and (2) implementing a "digestion-free" method by adding 5% Tween 20 to standard and sample preparations. The new method has been extensively tested with AAV of different serotypes, purification status, and transgenes encapsidated and was found to be highly accurate, precise, and robust. This significantly improved and simplified assay can be easily adopted by researchers in the gene therapy field and further automated for high-throughput applications.
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