Characterizing vaccines with light scattering

Importance of Topic

Vaccines play a central role in public health by preventing infectious diseases in humans and animals. Accurate characterization of vaccine components and carriers—including proteins, virus-like particles (VLPs), lipid nanoparticles (LNPs) and conjugates—is essential for discovery, formulation optimization, process control and regulatory compliance. Light scattering techniques provide label-free, absolute measurements of size, molar mass, charge and concentration that inform on stability, aggregation, molecular interactions and composition.

Objectives and Overview

This study reviews how static light scattering (multi-angle light scattering, MALS), dynamic light scattering (DLS) and electrophoretic light scattering (ELS) are applied to vaccine research and manufacturing. It describes the range of vaccine formats—from subunit proteins and polysaccharide conjugates to VLPs and nucleic acid-loaded nanoparticles—and illustrates how light scattering measurements guide key decision points in development and production.

Used Instrumentation

Key instruments and methods include:

• MALS detectors (DAWN, miniDAWN, microDAWN, UltraDAWN) paired with refractive index (Optilab) and UV concentration detectors
• Online dynamic light scattering modules (WyattQELS)
• Batch DLS instruments (DynaPro Plate Reader, NanoStar, ZetaStar)
• Electrophoretic light scattering for zeta potential (ZetaStar)
• Separation techniques: size-exclusion chromatography (SEC, UHPLC) and asymmetrical flow field-flow fractionation (FFF)
• Composition-gradient MALS (CG-MALS) for binding affinity and stoichiometry
• Real-time MALS (RT-MALS) for process analytical technology (PAT) using OBSERVER software

Main Results and Discussion

Case studies demonstrate how light scattering resolves critical vaccine attributes:

• Antibody-antigen interactions: SEC-MALS and CG-MALS define oligomeric state, stoichiometry and binding affinity of viral glycoprotein complexes
• Glycoprotein analysis: Combined UV and refractive index detection quantifies protein and glycan mass fractions in coronavirus spike antigens (~25% glycan by mass)
• Nucleic acid cargo in nanoparticles: Conjugate analysis distinguishes DNA or mRNA loading in VLPs and LNPs, revealing cargo per particle and empty versus full populations
• Size distribution: FFF-MALS (optionally with DLS) produces high-resolution radius distributions for LNPs and VLPs, crucial for delivery efficiency
• Stability and aggregation screening: Batch DLS and fractionation identify aggregation onset, thermal stress effects and optimal excipient formulations
• Zeta potential and charge: ELS paired with DLS distinguishes formulation variants and predicts colloidal stability and cellular uptake
• Process monitoring: RT-MALS measures molar mass, size and particle concentration in real time to automate fraction collection and ensure consistent product quality

Benefits and Practical Applications

Light scattering techniques enable:

• Absolute quantification of key quality attributes without labels
• Rapid, low-volume analysis suitable for high-throughput formulation screens
• Detection and sizing of multi-modal or aggregated populations
• Real-time monitoring for PAT and streamlined process control
• Flexible coupling to chromatography or FFF for advanced separation

Future Trends and Possible Uses

Emerging directions include:

• Integration of multi-detector platforms for simultaneous measurements of size, mass, charge and composition
• High-throughput screening of formulations and binding interactions using microfluidic and plate-based approaches
• Development of inline light scattering sensors for continuous biomanufacturing
• Advanced modeling to correlate in vitro light scattering data with in vivo distribution and immunogenicity
• Expansion of FFF-MALS to characterize novel nanoparticle carriers and complex biologics

Conclusion

Light scattering methods—MALS, DLS and ELS—offer a comprehensive toolkit for characterizing vaccines across discovery, development, manufacturing and quality control stages. Their ability to deliver absolute, label-free measurements of size, mass, charge and concentration improves understanding of molecular interactions, stability and composition. Coupled with separation techniques and real-time monitoring, these approaches support safe, efficient and reproducible vaccine production.

Reference

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