ArticleHuman gene therapy2021
Optimized Reversed-Phase Liquid Chromatography/Mass Spectrometry Methods for Intact Protein Analysis and Peptide Mapping of Adeno-Associated Virus Proteins.
Article in Human gene therapy, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 18 papers.
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Who cites it
18 citing papers in PubMed.
- Determination of AAV Capsid Protein Ratios by Sodium Dodecyl Sulfate Capillary Electrophoresis (CE-SDS)-UV Absorption Method.Electrophoresis · 2026Article
- Altering VP1 and VP2 expression in trans affects the transduction efficiency of AAV9.Frontiers in bioengineering and biotechnology · 2026Article
- Establishment of a novel human amniotic epithelial-derived cell line, HAT, for high-yield AAV vector production.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
- Combination of hydrophilic interaction liquid chromatography and top-down mass spectrometry for characterisation of adeno-associated virus capsid proteins.Analytical and bioanalytical chemistry · 2025Article
- Adeno-associated virus serotype 2 capsids with proteolytic cuts by trypsin remain intact and potent.Gene therapy · 2025Article
- A comparison of SWATH-MS methods for measurement of residual host cell proteins in adeno-associated virus preparations.Frontiers in bioengineering and biotechnology · 2025Article
- Quantitative proteomic analysis of residual host cell protein retention across adeno-associated virus affinity chromatography.Molecular therapy. Methods & clinical development · 2024Article
- Production of recombinant adeno-associated virus 5 using a novel self-attenuating adenovirus production platform.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
- Assessment of adeno-associated virus purity by capillary electrophoresis-based western.Molecular therapy. Methods & clinical development · 2024Article
- Characterizing Glycosylation of Adeno-Associated Virus Serotype 9 Capsid Proteins Generated from HEK293 Cells through Glycopeptide Mapping and Released Glycan Analysis.Microorganisms · 2024Article
- Development of a Rapid Adeno-Associated Virus (AAV) Identity Testing Platform through Comprehensive Intact Mass Analysis of Full-Length AAV Capsid Proteins.Journal of proteome research · 2023Article
- Correlating physicochemical and biological properties to define critical quality attributes of a rAAV vaccine candidate.Molecular therapy. Methods & clinical development · 2023Article
- Development of an icIEF assay for monitoring AAV capsid proteins and application to gene therapy products.Molecular therapy. Methods & clinical development · 2023Article
- Endothelial cell-derived MMP19 promotes pulmonary fibrosis by inducing E(nd)MT and monocyte infiltration.Cell communication and signaling : CCS · 2023Article
- Fast and high-throughput LC-MS characterization, and peptide mapping of engineered AAV capsids using LC-MS/MS.Molecular therapy. Methods & clinical development · 2022Article
- Assessing production variability in empty and filled adeno-associated viruses by single molecule mass analyses.Molecular therapy. Methods & clinical development · 2022Article
Corrections and comments
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Authors and funding
7 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Recombinant adeno-associated viruses (AAVs) have emerged as the leading gene delivery platform owing to their nonpathogenic nature and long-term gene expression capability. The AAV capsid, in addition to protecting the viral genome, plays an important role in viral infectivity and gene transduction, indicating the value of the constituent viral proteins (VPs) being well-characterized as part of gene therapy development. However, the limited sample availability and sequence homology shared by the VPs pose challenges to adapt existing analytical methods developed for conventional biologics. In this study, we report the development of reversed-phase liquid chromatography/mass spectrometry-based methods for characterization of AAV capsid proteins at intact protein and peptide level with reduced sample consumptions. The developed methods allowed the measurement of VP expression with fluorescence detection and intact mass/post-translational modifications (PTMs) analysis through a benchtop time-of-flight mass spectrometer. The general applicability and validity of the methods for gene therapy product development were demonstrated by applying the optimized methods to multiple common AAV serotypes. A 1-h enzymatic digestion method was also developed using 1.25 μg of AAV VPs, providing >98% protein sequence coverage and reproducible relative quantification of various PTMs of the VPs. The efficient and sensitive analyses of AAV capsid proteins enabled by the reported methods provide further understanding and offer guidance in the development and manufacturing of AAV-related therapeutics.
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