Fusion-dependent formation of lipid nanoparticles containing macromolecular payloads

Jayesh A. Kulkarni (Corresponding author), Dominik Witzigmann, Jerry Leung, Roy van der Meel, Josh Zaifman, Maria M. Darjuan, Hiu Man Grisch-Chan, Beat Thöny, Yuen Yi C. Tam, Pieter R. Cullis (Corresponding author)

Research output: Contribution to journalArticleAcademicpeer-review

82 Citations (Scopus)
230 Downloads (Pure)


The success of Onpattro™ (patisiran) clearly demonstrates the utility of lipid nanoparticle (LNP) systems for enabling gene therapies. These systems are composed of ionizable cationic lipids, phospholipid, cholesterol, and polyethylene glycol (PEG)-lipids, and are produced through rapid-mixing of an ethanolic-lipid solution with an acidic aqueous solution followed by dialysis into neutralizing buffer. A detailed understanding of the mechanism of LNP formation is crucial to improving LNP design. Here we use cryogenic transmission electron microscopy and fluorescence techniques to further demonstrate that LNP are formed through the fusion of precursor, pH-sensitive liposomes into large electron-dense core structures as the pH is neutralized. Next, we show that the fusion process is limited by the accumulation of PEG-lipid on the emerging particle. Finally, we show that the fusion-dependent mechanism of formation also applies to LNP containing macromolecular payloads including mRNA, DNA vectors, and gold nanoparticles.
Original languageEnglish
Pages (from-to)9023-9031
Number of pages9
Issue number18
Early online date22 Apr 2019
Publication statusPublished - 14 May 2019


  • Cryoelectron Microscopy
  • Genetic Therapy/methods
  • Hydrogen-Ion Concentration
  • Lipids/chemistry
  • Liposomes/chemistry
  • Macromolecular Substances/chemistry
  • Nanoparticles/chemistry
  • Particle Size
  • Polyethylene Glycols/chemistry
  • RNA, Messenger/chemistry
  • RNA, Small Interfering/chemistry


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