Fig. 2: In vitro LNP-mediated pDNA delivery.

a Transfection efficiency of LNPs in HepG2 cells via high-throughput screening platform after 72 h incubation (n = 2). The level of transgene expression for each formulation is shown using luciferase as a reporter. b The top 32 formulations from each helper lipid series were selected based on transfection efficiency in HepG2. FACS was used to further evaluate the transfection efficiency of c DOTAP, d DOPE and e 18PG series of LNPs using GFP as a reporter gene at a pDNA dose of 1 μg/mL; gene expression was analyzed at 72 h after transfection (n = 2). Cellular metabolic activity was measured by alamarBlue assay (n = 4). Formulations were regrouped into four clusters, each containing eight formulations, based on their transgene expression level. Data are presented as mean±S.D. The percentage of each component in the formulations is indicated by pie charts: Ionizable lipid (red), cholesterol (green), helper lipid (blue), DMG-PEG2000 (yellow). See Supplementary Tables 1–6 for molar percentage used in the 32 formulations for each helper lipid. Bars refer to the MFI (Median fluorescence intensity) value on the left, the boxes refer to the metabolic activity on the right. f Histogram of the Z-average diameters of top 32 LNP formulations made from each helper lipid. g Percentage of LNP formulations with size less than 200 nm and less than 400 nm for each helper lipid. Source data are provided as a Source Data file.