Key publications on the characterization of nanoparticle drug and gene delivery systems by FFF-MALS-DLS

Importance of the Topic

Nanoparticle-based drug and gene delivery systems demand rigorous physicochemical characterization to guarantee performance, safety, and regulatory acceptance. Asymmetric-flow field-flow fractionation coupled with multi-angle light scattering and dynamic light scattering (AF4-MALS-DLS) provides a versatile, high-resolution platform to measure particle size distributions, molecular weight, concentration, drug loading, and stability across a wide range of nanocarriers.

Objective and Study Overview

This collection presents pivotal publications that apply AF4-MALS-DLS to characterize nanoDDS, including lipid nanoparticles, liposomes, exosomes, polymersomes, virus-like particles, and viral vectors. The goal is to illustrate methodological developments, cross-validation with orthogonal techniques, and case studies demonstrating the approach’s value in preclinical and quality control settings.

Methodology and Instrumentation

• Separation Principle: AF4 employs a crossflow to gently fractionate particles by size.
• Detectors: MALS yields radius and molar mass; DLS provides hydrodynamic size; UV and refractive index detectors offer concentration and compositional data.
• Workflow: Stepwise increase in sample complexity, from standard beads to clinically relevant formulations, combining AF4-MALS-DLS with complementary assays (TEM, mass spectrometry, fluorescence).

Key Results and Discussion

• Lipid Carriers: Optimized AF4 protocols quantified size, polydispersity, and drug release kinetics of liposomes and lipid nanoparticles for mRNA delivery.
• Exosomes and Extracellular Vesicles: AF4 enabled high-resolution separation of vesicle subpopulations, supporting lipidomic profiling and biomarker discovery.
• Polymersomes: Studies elucidated membrane permeability and enzyme diffusion through synthetic vesicle membranes.
• Virus-Like Particles and Viral Vectors: AF4-MALS-DLS provided detailed size distributions, aggregation state, and assembly quality for vaccine candidates and gene therapy vehicles.
• Other Modalities: Application to metal–organic frameworks and dendrimer conjugates highlighted broad utility in nanomedicine.

Benefits and Practical Applications

• Non-invasive, gentle fractionation preserves particle integrity.
• High-resolution sizing supports detection of subtle differences in formulation batches.
• Quantitative analysis of drug loading and release kinetics guides formulation optimization.
• Regulatory relevance through validated, orthogonal characterization workflows.

Future Trends and Potential Applications

Emerging directions include integration of microfluidic FFF systems for higher throughput, coupling with mass spectrometry for compositional analysis, application of machine learning for data interpretation, and expanded use in personalized nanomedicine and real-time process monitoring.

Conclusion

AF4-MALS-DLS has become an indispensable analytical tool in the development and quality control of nanoparticle-based therapeutics. Its ability to resolve complex mixtures, quantify critical quality attributes, and interface with orthogonal methods drives innovation in drug delivery research and supports regulatory compliance.

References

Key references include Caputo et al. Mol. Pharm. 16(2):756–767 (2019); Mehn et al. EUNCL-PCC-022 (2016); Mildner et al. Eur. J. Pharm. Biopharm. 163:252–265 (2021); Parot et al. J. Controlled Release 320:495–510 (2020); Wu et al. Anal. Chim. Acta 1127:234–245 (2020); Zhang H. Nat. Protoc. 14(4):1–18 (2019); Petersen et al. Anal. Bioanal. Chem. 406(30):7855–7866 (2014); Bousse et al. J. Virol. Methods 193(2):589–596 (2013); Roda et al. Anal. Bioanal. Chem. 410(21):5245–5253 (2018); Sonzini et al. Int. J. Pharm. 637:122905 (2023).