A deep-ultraviolet Raman spectroscopy approach may offer a new way to evaluate how messenger RNA (mRNA) is packaged inside lipid nanoparticles (LNPs), a key step in the development of mRNA vaccines and therapeutics.
Researchers at the University at Albany, SUNY demonstrated that the technique can distinguish between encapsulated and free mRNA molecules within nanoparticle formulations. Because lipid nanoparticles protect fragile RNA molecules and deliver them into cells, assessing encapsulation efficiency is a key step in formulation and quality control.
“mRNA therapeutics have emerged as a powerful tool for treating a wide range of diseases, but their clinical success depends on overcoming issues of instability and delivery,” said Igor Lednev, University at Albany, who led the study. “Raman spectroscopy offers us unique information that can help ensure mRNA is fully encapsulated inside lipid nanoparticles, ensuring the safety and effectiveness of these therapeutics.”
In the study, the researchers used deep-UV resonance Raman (DUVRR) spectroscopy, which measures molecular vibrations by detecting how laser light scatters from a sample. Using a deep-UV excitation wavelength of 266 nm – close to the absorption maximum of nucleic acids – the method selectively enhances Raman signals from the mRNA while minimizing interference from surrounding lipids.
The team prepared model vaccine formulations containing different ratios of mRNA and lipid nanoparticles, generating samples with fully encapsulated, partially encapsulated, and free RNA molecules. Analysis of the resulting spectra revealed distinct vibrational markers associated with lipid–mRNA interactions. In particular, a Raman band near 1322 cm⁻¹ was linked to lipid-bound mRNA, while a band near 1329 cm⁻¹ corresponded to free RNA molecules.
To interpret these spectral differences, the team combined Raman measurements with principal component analysis and two-dimensional correlation spectroscopy. The analysis showed that changes in the 1322 cm⁻¹ signal tracked the degree of mRNA encapsulation, suggesting it could serve as a quantitative indicator of nanoparticle formulation efficiency. The study also applied a two-trace two-dimensional correlation spectroscopy (2T2D-COS) approach to isolate spectral contributions linked specifically to lipid–mRNA interactions.
“Raman spectroscopy allows us to analyze mRNA inside lipid nanoparticles without damaging it,” said Alexander Shekhtman, a collaborator on the study. “This means we can optimize formulations to improve both safety and effectiveness.”
The researchers suggest that the approach could eventually support formulation screening and quality control for mRNA-based therapeutics, offering a nondestructive way to monitor encapsulation during development and manufacturing.
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