Multiparametric Characterization of Individual Suspended Nanoparticles Using Confocal Fluorescence and...

Area of Science:

• Nanotechnology
• Biophysics
• Materials Science

Background:

• Accurate characterization of mRNA-lipid nanoparticles (LNPs) is crucial for understanding their therapeutic efficacy.
• Current methods often provide only ensemble-average properties or require specialized equipment.
• Limitations in characterizing individual LNP properties hinder therapeutic development.

Purpose of the Study:

• To develop a quantitative, in-solution method for simultaneous characterization of individual LNPs.
• To overcome limitations of existing ensemble-average or specialized microscope-dependent methods.
• To enable widespread adoption of advanced nanoparticle characterization.

Main Methods:

• Combined a commercially available confocal microscope with a convex lens-induced confinement (CLiC) instrument.
• Achieved simultaneous characterization of size, mass, refractive index, and nucleic acid payload of individual LNPs.
• Validated the method using nanosized beads and investigated LNPs in varying solvent pH.

Main Results:

• Successfully correlated size, mass, refractive index, and nucleic acid payload of individual LNPs.
• Demonstrated the accuracy and precision of the developed method.
• Investigated the influence of solvent pH on LNP size, payload, and water content.

Conclusions:

• The developed method provides a accessible approach for detailed LNP characterization.
• This technique facilitates a deeper understanding of LNP heterogeneity and its impact on therapeutic outcomes.
• The use of readily available microscopy tools promotes broader application in nanoparticle research and development.