Time-Resolved and Label-Free Evanescent Light-Scattering Microscopy for Mass Quantification of Protein Binding to Single Lipid Vesicles.

Mattias Sjöberg,Fredrik Höök,Björn Agnarsson,Mokhtar Mapar,Antonius Armanious,Vladimir P Zhdanov

doi:10.1021/acs.nanolett.1c00644

Mattias Sjöberg, Fredrik Höök + Show 4 more

Open Access

https://doi.org/10.1021/acs.nanolett.1c00644

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Abstract

In-depth understanding of the intricate interactions between biomolecules and nanoparticles is hampered by a lack of analytical methods providing quantitative information about binding kinetics. Herein, we demonstrate how label-free evanescent light-scattering microscopy can be used to temporally resolve specific protein binding to individual surface-bound (∼100 nm) lipid vesicles. A theoretical model is proposed that translates protein-induced changes in light-scattering intensity into bound mass. Since the analysis is centered on individual lipid vesicles, the signal from nonspecific protein binding to the surrounding surface is completely avoided, offering a key advantage over conventional surface-based techniques. Further, by averaging the intensities from less than 2000 lipid vesicles, the sensitivity is shown to increase by orders of magnitude. Taken together, these features provide a new avenue in studies of protein-nanoparticle interaction, in general, and specifically in the context of nanoparticles in medical diagnostics and drug delivery.

Highlights

In-depth understanding of the intricate interactions between biomolecules and nanoparticles is hampered by a lack of analytical methods providing quantitative information about binding kinetics
Nanoparticle tracking analysis (NTA)[7] and flow cytometry (FC)[8] provide high-quality statistics but are restricted to NP size determination, in the case of NTA, and subpopulation identification using fluorescent markers in the case of FC, and neither approach is well suited for investigating kinetics
Considering the fact that biological NPs often have broad distributions in terms of composition, structure, and size, and that the associated biomolecular interaction kinetics strongly depends on such properties and may exhibit features that are hidden at the ensemble level,[12] there is a need for methods that operate at the single-NP level to enable investigation of sample heterogeneity, while simultaneously providing label-free quantitative readout with sufficient statistics and temporal resolution to enable investigations of interaction kinetics

Summary

■ REFERENCES

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■ ACKNOWLEDGMENTS