ArticlePharmaceutics2021
Multiple-Monitor HPLC Assays for Rapid Process Development, In-Process Monitoring, and Validation of AAV Production and Purification.
Article in Pharmaceutics, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 14 papers.
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Who cites it
14 citing papers in PubMed, 42 citations in OpenAlex.
- Mechanistic Insights into AAV Capsid-Stationary Phase Interactions Governing Native Stability and Chromatographic Separation Using AAV8 as a Model System.Pharmaceutics · 2026Article
- A two-pass anion-exchange chromatography strategy for enrichment of full capsids in manufacturing of adeno-associated viral vectors.Molecular therapy. Methods & clinical development · 2025Article
- Chromatography in Downstream Processing of Recombinant Adeno-Associated Viruses: A Review of Current and Future Practises.Biotechnology and bioengineering · 2025Review
- Model-Based Characterization of the Metabolism of Recombinant Adeno-Associated Virus (rAAV) Production via Human Embryonic Kidney (HEK293) Cells.Bioengineering (Basel, Switzerland) · 2025Article
- Application of aqueous two-phase extraction for separation and purification of various adeno-associated viruses.Biotechnology letters · 2025Article
- Size-exclusion chromatography as a multi-attribute method for process and product characterization of adeno-associated virus.Molecular therapy. Methods & clinical development · 2024Article
- Quantification of full and empty particles of adeno-associated virus vectors via a novel dual fluorescence-linked immunosorbent assay.Molecular therapy. Methods & clinical development · 2024Article
- Evaluation of a rapid multi-attribute combinatorial high-throughput UV-Vis/DLS/SLS analytical platform for rAAV quantification and characterization.Molecular therapy. Methods & clinical development · 2024Article
- Adeno-associated viral capsid stability on anion exchange chromatography column and its impact on empty and full capsid separation.Molecular therapy. Methods & clinical development · 2023Article
- Comprehensive Comparison of AAV Purification Methods: Iodixanol Gradient Centrifugation vs. Immuno-Affinity Chromatography.Advances in cell and gene therapy · 2023Article
- Biophysical Characterization of Adeno-Associated Virus Vectors Using Ion-Exchange Chromatography Coupled to Light Scattering Detectors.International journal of molecular sciences · 2022Article
- Reverse-Phase Ultra-Performance Chromatography Method for Oncolytic Coxsackievirus Viral Protein Separation and Empty to Full Capsid Quantification.Human gene therapy · 2022Article
- Article
- Anion-exchange HPLC assay for separation and quantification of empty and full capsids in multiple adeno-associated virus serotypes.Molecular therapy. Methods & clinical development · 2021Article
Corrections and comments
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Authors and funding
7 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
HPLC is established as a fast convenient analytical technology for characterizing the content of empty and full capsids in purified samples containing adeno-associated virus (AAV). UV-based monitoring unfortunately over-estimates the proportion of full capsids and offers little value for characterizing unpurified samples. The present study combines dual-wavelength UV monitoring with intrinsic fluorescence, extrinsic fluorescence, and light-scattering to extend the utility of HPLC for supporting development of therapeutic AAV-based drugs. Applications with anion exchange (AEC), cation exchange (CEC), and size exclusion chromatography (SEC) are presented. Intrinsic fluorescence increases sensitivity of AAV detection over UV and enables more objective estimation of empty and full capsid ratios by comparison of their respective peak areas. Light scattering enables identification of AAV capsids in complex samples, plus semiquantitative estimation of empty and full capsid ratios from relative peak areas of empty and full capsids. Extrinsic Picogreen fluorescence enables semiquantitative tracking of DNA with all HPLC methods at all stages of purification. It does not detect encapsidated DNA but reveals DNA associated principally with the exteriors of empty capsids. It also enables monitoring of host DNA contamination across chromatograms. These enhancements support many opportunities to improve characterization of raw materials and process intermediates, to accelerate process development, provide rapid in-process monitoring, and support process validation.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.