Snag spot-on adenovirus titer & characterization with Stunner

Importance of the topic

Gene therapy and vaccine applications depend on precise measurement of adenoviral vector concentration and empty/full capsid ratio. Rapid, reliable analytics accelerate process development, quality control, and ensure consistent product efficacy.

Aims and overview of the study

This application note evaluates the Unchained Labs Stunner platform with its Adeno Quant application. By integrating UV/Vis spectroscopy, dynamic light scattering (DLS), and static light scattering (SLS), it delivers comprehensive adenovirus 5 (AdV-5) characterization—capsid titer, empty/full ratio, protein/DNA quantification, size, and aggregation—without sample denaturation or extensive reagents.

Methodology and Instrumentation

Three lots of CsCl-purified AdV-5 from three vendors were thawed once, stored at 4 °C, and measured within four days. Each 2 µL sample underwent four 5 s DLS acquisitions, full-spectrum UV/Vis scanning, and SLS intensity measurement. Unmix algorithms corrected for turbidity and separated absorbance contributions from protein, dsDNA, and light scattering. Molar extinction coefficients were calculated from published amino acid sequences and genome sizes. The Adeno Quant application outputs total capsid titer, full capsid titer, free and aggregated protein and DNA titers, percent empty/full, Z-average size, and polydispersity index (PDI).

Used Instrumentation

• Stunner platform (Unchained Labs) combining UV/Vis, DLS, and SLS modules
• Adeno Quant software with built-in calculations and optional 21 CFR Part 11 compliance
• 96-well autosampler capacity; 2 µL sample volume per measurement

Main results and discussion

Vendor 1 samples displayed high purity: total capsids ~4.2 × 10^12 cp/mL (CV 4.9%), full capsids ~3.4 × 10^12 vg/mL (CV 4.4%), 81% full, monodisperse size distribution, and ~13% free/aggregated protein and DNA. Vendor 2 showed correct hydrodynamic size (Z-average 122.6 nm, PDI < 0.1) but excess dsDNA, indicating host cell DNA contamination. Vendor 3 had appropriate protein and DNA mass but significant aggregation detected by DLS, implying stability issues. Integrating UV/Vis with light scattering data distinguished true capsid signal from contaminants, revealing sample-specific quality attributes.

Benefits and practical applications

The Stunner Adeno Quant workflow is fast (≤ 1 h for 96 samples), label-free, serotype-agnostic, and requires minimal sample preparation. Small volume requirements and robust turbidity correction enable at-line process monitoring, KPI/CQA development, and expedited decision-making in research and GMP environments.

Future trends and possibilities for use

• Full automation and expanded throughput for high-density plate formats
• Extension to other viral vectors and nanoparticle platforms in gene therapy and vaccine research
• Integration with process analytical technology (PAT) frameworks and advanced data analytics for real-time monitoring and control

Conclusion

By uniting UV/Vis, DLS, and SLS in one platform, Stunner’s Adeno Quant application provides rapid, multiparametric adenovirus vector analysis without complex sample handling. This streamlined approach supports accelerated process development, robust quality control, and reliable assessment of titer, empty/full ratio, size, and aggregation.

References

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