Electrophoretic Microfluidic Characterization of mRNA- and pDNA-Loaded Lipid Nanoparticles

Area of Science:

• Biotechnology
• Nanotechnology
• Analytical Chemistry

Background:

• Lipid nanoparticles (LNPs) are advanced nonviral gene delivery systems for treating viral, oncological, and genetic diseases.
• Developing LNP therapies necessitates rapid analytical methods for efficient development and manufacturing.
• Current analytical techniques can be time-consuming, limiting high-throughput screening.

Purpose of the Study:

• To develop and validate a rapid, high-throughput microfluidic electrophoresis method for analyzing LNP composition.
• To enable accurate quantification of lipid and nucleic acid content within LNPs for optimized therapeutic loading.
• To support the research, development, and manufacturing of LNP-based gene therapies.

Main Methods:

• Utilized an electrophoresis microfluidic platform for LNP analysis.
• Analyzed LNPs with varying clinical lipid compositions (Onpattro, Comirnaty, Spikevax) and nucleic acids (plasmid DNA, mRNA).
• Employed a 96- or 384-well plate format for high-throughput screening, with sample analysis times of 2-4 minutes and a volume of 11 μL.

Main Results:

• Achieved rapid analysis of LNP lipid and nucleic acid formulations with high precision.
• Lipid analysis showed average precision error of 10.4% and prediction error of 19.1% compared to NanoSight.
• Nucleic acid analysis demonstrated average precision error of 4.8% and prediction error of 9.4% compared to PicoGreen/RiboGreen assays.
• Quantified nucleic acid per LNP, finding an average of 263 mRNA/LNP and 126 pDNA/LNP.

Conclusions:

• The developed microfluidic electrophoresis method offers a rapid and accurate approach for LNP characterization.
• This technique facilitates high-throughput screening, crucial for LNP manufacturing and formulation R&D.
• The method's ability to quantify nucleic acid per LNP aids in optimizing therapeutic loading and efficacy.