Payload distribution and capacity of mRNA lipid nanoparticles

Lipid nanoparticles (LNPs) are effective vehicles to deliver mRNA vaccines and therapeutics. It has been challenging to assess mRNA packaging characteristics in LNPs, including payload distribution and capacity, which are critical to understanding structure-property-function relationships for furthe...

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Bibliographic Details
Published in:Nature communications 2022-09, Vol.13 (1), p.5561-5561, Article 5561
Main Authors: Li, Sixuan, Hu, Yizong, Li, Andrew, Lin, Jinghan, Hsieh, Kuangwen, Schneiderman, Zachary, Zhang, Pengfei, Zhu, Yining, Qiu, Chenhu, Kokkoli, Efrosini, Wang, Tza-Huei, Mao, Hai-Quan
Format: Article
Language:English
Subjects:
639/301/54/152
639/925/352
639/925/930/12
Assembly
Fluorescence
Humanities and Social Sciences
Illumination
Lipids
mRNA
mRNA vaccines
multidisciplinary
Nanoparticles
Packaging
Science
Science (multidisciplinary)
Spectroscopy
Spectrum analysis
Structure-function relationships
Vaccines
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Summary:Lipid nanoparticles (LNPs) are effective vehicles to deliver mRNA vaccines and therapeutics. It has been challenging to assess mRNA packaging characteristics in LNPs, including payload distribution and capacity, which are critical to understanding structure-property-function relationships for further carrier development. Here, we report a method based on the multi-laser cylindrical illumination confocal spectroscopy (CICS) technique to examine mRNA and lipid contents in LNP formulations at the single-nanoparticle level. By differentiating unencapsulated mRNAs, empty LNPs and mRNA-loaded LNPs via coincidence analysis of fluorescent tags on different LNP components, and quantitatively resolving single-mRNA fluorescence, we reveal that a commonly referenced benchmark formulation using DLin-MC3 as the ionizable lipid contains mostly 2 mRNAs per loaded LNP with a presence of 40%–80% empty LNPs depending on the assembly conditions. Systematic analysis of different formulations with control variables reveals a kinetically controlled assembly mechanism that governs the payload distribution and capacity in LNPs. These results form the foundation for a holistic understanding of the molecular assembly of mRNA LNPs. Lipid nanoparticles (LNPs) are effective vehicles to deliver mRNA vaccines and therapeutics but assessing the mRNA packaging characteristics in LNPs is challenging. Here, the authors report that mRNA and lipid contents in LNP formulations can be quantitatively examined by multi-laser cylindrical illumination confocal spectroscopy at the single-nanoparticle level.
ISSN:2041-1723
2041-1723
DOI:10.1038/s41467-022-33157-4