Hassle-free nanoparticle characterization with Stunner

Importance of the Topic

Nanoparticle‐based carriers are transforming drug delivery, gene therapy and vaccine research. Precise determination of payload concentration and particle size is essential to predict therapeutic efficacy and biodistribution. Conventional assays often rely on disruptive workflows, fluorescent dyes or complex chromatography, hindering throughput and introducing variability.

Objectives and Overview

This application note illustrates how the Stunner platform streamlines nanoparticle characterization by merging UV/Vis absorbance deconvolution with dynamic light scattering (DLS). It demonstrates non‐destructive quantification of RNA, DNA, proteins and small‐molecule payloads, simultaneous size measurement and lipid absorbance analysis in a unified, high‐throughput workflow.

Methodology and Instrumentation

• Lipid nanoparticles (LNPs) containing RNA, DNA or lysozyme were prepared using a microfluidic mixer (NanoAssemblr Ignite) at defined N/P ratios and lipid compositions (DOTAP:POPE:Chol:DMG‐PEG).
• Samples were buffer‐exchanged into PBS and analyzed with Stunner using 2 µL per well in SBS‐format plates.
• UV/Vis spectra were deconvoluted with Unmix algorithms to separate payload, turbidity and lipid/buffer contributions. Protein quantification used absorbance at 280 nm and extinction coefficients.
• Hydrodynamic diameter and polydispersity index (PDI) were measured by DLS (5 s acquisitions, 4 °C), validated with gold nanoparticle standards.
• The Custom Nanoparticle app utilized stored analyte spectra to quantify doxorubicin content in PEGylated liposomes.

Main Results and Discussion

• RNA, DNA and protein LNPs exhibited highly linear quantification over dilution series (R²>0.99, CV<5%).
• Lipid/buffer absorbance at 230 nm also correlated linearly with sample dilution (R²>0.92).
• Particle size varied with N/P ratio: higher nucleic acid loading yielded larger diameters, while PDI remained below 0.2 for most samples.
• Doxorubicin quantification matched target concentrations (slope ~1, R²>0.99), and mixed liposome populations showed predictable size and polydispersity changes.

Benefits and Practical Applications

• Non‐destructive, reagent‐free payload quantification eliminates detergents, dyes and standard curves.
• Low sample volume (2 µL) and 96‐well throughput enable rapid screening of formulations in under one hour.
• Combined UV/Vis and DLS measurements facilitate integrated assessment of concentration, size and polydispersity for QA/QC, process development and stability studies.

Future Trends and Opportunities

• Expansion to novel nanoparticle types via user‐defined spectra and machine learning–driven unmixing.
• Integration into automated platforms and compliance with 21 CFR Part 11 for regulated environments.
• Real‐time monitoring of manufacturing streams and in‐line quality control in biopharmaceutical production.

Conclusion

The Stunner platform offers a unique, high‐throughput solution for comprehensive nanoparticle characterization. By combining UV/Vis deconvolution with DLS sizing, it delivers accurate payload quantification and size measurements in a single, non‐destructive step, accelerating R&D and quality workflows across gene therapy, vaccine and drug delivery applications.

References

1. Roces C. et al., Pharmaceutics, 2016;8(3): Rapid quantification and validation of lipid concentrations within liposomes.
2. Roces C. et al., Pharmaceutics, 2020;12(11): Manufacturing considerations for lipid nanoparticles using microfluidics.
3. Ribeiro L. et al., Scientific Reports, 2018;8(1): Use of nanoparticle concentration to understand colloid structure.
4. Hussain M. et al., Pharmaceutics, 2019;11(1): Comparative analysis of protein quantification methods in liposomal formulations.
5. Duong V. et al., Molecules, 2020;25(20):4781. Preparation of solid lipid nanoparticles and nanostructured lipid carriers by solvent injection.