MembranesJun 25, 2026

Real-Time Tracking of Lipid Nanoparticles Forming Using Time-Resolved X-Ray Scattering

mRNA Technology Weekly Brief ↗PubMed ↗DOI ↗

Updated Jun 26, 2026

Abstract

A time-resolved in situ small-angle X-ray scattering method was developed to monitor the structural evolution of lipid nanoparticles during formulation.

Distinct assembly pathways were observed for empty lipid nanoparticles (LNPs) and those encapsulated with messenger RNA (mRNA).
Empty-LNPs transitioned gradually toward periodic nanostructures.
mRNA-LNPs complexed rapidly into stable subunits, followed by hierarchical assembly.
Nanoscale structural rearrangements during microfluidic dilution were tracked, indicated by shifts in scattering peaks and internal periodicity.
The methodology captures transient intermediate states, aiding in understanding molecular assembly mechanisms.

Simplified

Lipid nanoparticles (LNPs) have emerged as popular nucleic acid delivery systems, yet the dynamic mechanisms related to their self-assembly and structural maturation remain insufficiently understood due to the limitations of traditional offline characterization tools. This study establishes a time-resolved (TR) in situ small-angle X-ray scattering (SAXS) methodology to monitor the structural evolution of LNPs during microfluidic formulation and subsequent maturation. By integrating a dual-channel microfluidic mixing system with a SAXS measurement platform, we successfully captured the real-time scattering profiles of both empty and messenger RNA-loaded nanoparticles (mRNA-LNPs). The results demonstrate distinct assembly pathways for empty-LNPs and those encapsulated with mRNA. The empty-LNPs undergo a gradual transition toward periodic nanostructures, whereas mRNA-LNPs exhibit rapid complexation into stable subunits followed by hierarchical assembly. Furthermore, the platform effectively tracked nanoscale structural rearrangements during a microfluidic dilution process, revealed by subtle shifts in scattering peaks and internal periodicity. Overall, this time-resolved approach provides a robust experimental framework for capturing transient intermediate states, offering a valuable tool to elucidate molecular assembly mechanisms and facilitate the rational design of next-generation nanomedicines.

Funding

Competing interests

The authors declare no conflicts of interest.

PubMed

Real-Time Tracking of Lipid Nanoparticles Forming Using Time-Resolved X-Ray Scattering

Abstract

Funding

Competing interests

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