Proven Intelligence Accelerating Next-Generation Therapies.

Executive Summary

Confirming the precise stoichiometric ratio of the three adeno-associated virus (AAV) capsid viral proteins (VP1, VP2, and VP3) is a required analytical step for product characterization and lot release. Intact mass analysis via liquid chromatography-mass spectrometry (LC-MS) provides definitive data on protein identity, integrity, and the molar ratio of these subunits. This analysis directly informs product consistency, which is linked to vector potency, stability, and immunogenicity, forming a key component of the data package for Investigational Medicinal Product Dossier (IMPD) submissions, particularly for sponsors targeting Swissmedic or other European agencies.

Verifying AAV Capsid Stoichiometry

The assembly of an AAV capsid from its constituent viral proteins (VP1, VP2, and VP3) is a defined aspect of vector biology. The precise ratio of these proteins is a defined feature of the specific AAV serotype, directly linked to the final vector’s biological function. Inconsistent stoichiometry between manufacturing lots can introduce significant variability, impacting program timelines and regulatory acceptance.

Our analytical services provide definitive characterization of the VP1:VP2:VP3 ratio using intact mass analysis. This approach moves beyond simple gel-based estimations to deliver quantitative data, a requirement for robust CMC packages that support an accelerated 18-24 month path to IND.

Our approach is founded on a collaborative partnership model, providing sponsors with direct access to scientific teams with comprehensive expertise in vector production and analytics. This ensures clear communication and alignment on analytical strategies critical for program advancement.

The Link Between Capsid Composition and Biological Activity

The protein composition of the AAV capsid surface has direct consequences for therapeutic outcomes. Published data shows that specific capsid structures can influence the host immune response, with certain protein domains driving T cell activation (PMID: 19414756). This underscores the need to confirm that the manufactured vector’s capsid composition is consistent and matches the intended design, minimizing the risk of unintended immunogenicity.

The capsid assembly also directly impacts vector functionality. The efficiency of vector transduction into target cells is influenced by the capsid proteins present (PMID: 12829835). By verifying the correct VP ratio, sponsors gain higher confidence in the vector’s expected tropism and potency, which are key measures of therapeutic efficacy.

Our Analytical Facilities

Our >100,000 sq ft facility is equipped with advanced instrumentation to support comprehensive analytics for viral vectors.

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A GxP-Compliant Workflow for AAV Characterization

Our process for intact mass analysis is designed for accuracy and regulatory compliance, supporting programs from preclinical research through to IMPD submissions. While the Franklin Biolabs brand was launched in 2024, our scientific leadership’s track record has contributed to a 100% successful IND rate for client programs since 2019.

• Sample Preparation: The AAV sample undergoes denaturation to dissociate the capsid into its individual VP1, VP2, and VP3 subunits.

• Chromatographic Separation: Reversed-phase liquid chromatography (RPLC) is used to separate the three viral proteins based on their hydrophobicity.

• Mass Spectrometry Analysis: The separated proteins are introduced into a high-resolution mass spectrometer to determine the precise molecular weight of each intact protein.

• Data Deconvolution and Analysis: Specialized software deconvolutes the mass spectra to confirm the identity of each protein against its theoretical mass. The relative abundance of each protein is calculated from the chromatographic peak areas to determine the final stoichiometric ratio.

• Reporting: A comprehensive report is generated detailing the methodology, results, and data interpretation, suitable for inclusion in regulatory filings.

This workflow provides accelerated characterization for rapid product validation, delivering a clear, data-driven assessment of AAV capsid integrity. For more on our comprehensive analytical capabilities, please visit our parent hub page on Analytics.