Microfluidic Formulation of DNA-Loaded Multicomponent Lipid Nanoparticles for Gene Delivery

Erica Quagliarini, Serena Renzi, Luca Digiacomo, Francesca Giulimondi, Barbara Sartori, Heinz Amenitsch, Valentina Tassinari, Laura Masuelli, Roberto Bei, Lishan Cui, Junbiao Wang, Augusto Amici, Cristina Marchini, Daniela Pozzi*, Giulio Caracciolo*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


In recent years, lipid nanoparticles (LNPs) have gained considerable attention in numerous research fields ranging from gene therapy to cancer immunotherapy and DNA vaccination. While some RNA-encapsulating LNP formulations passed clinical trials, DNA-loaded LNPs have been only marginally explored so far. To fulfil this gap, herein we investigated the effect of several factors influencing the microfluidic formulation and transfection behavior of DNA-loaded LNPs such as PEGylation, total flow rate (TFR), concentration and particle density at the cell surface. We show that PEGylation and post-synthesis sample concentration facilitated formulation of homogeneous and small size LNPs with high transfection efficiency and minor, if any, cytotoxicity on human Embryonic Kidney293 (HEK-293), spontaneously immortalized human keratinocytes (HaCaT), immortalized keratinocytes (N/TERT) generated from the transduction of human primary keratinocytes, and epidermoid cervical cancer (CaSki) cell lines. On the other side, increasing TFR had a detrimental effect both on the physicochemical properties and transfection properties of LNPs. Lastly, the effect of particle concentration at the cell surface on the transfection efficiency (TE) and cell viability was largely dependent on the cell line, suggesting that its case-by-case optimization would be necessary. Overall, we demonstrate that fine tuning formulation and microfluidic parameters is a vital step for the generation of highly efficient DNA-loaded LNPs.

Original languageEnglish
Article number1292
Issue number8
Publication statusPublished - 19 Aug 2021


  • lipid nanoparticles
  • microfluidics
  • transfection efficiency
  • lipofectamine
  • Lipofectamine
  • Transfection efficiency
  • Lipid nanoparticles
  • Microfluidics

ASJC Scopus subject areas

  • Pharmaceutical Science

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