Fig. 6: Biodistribution, cellular uptake and endosomal escape levels of top-performing LNP formulations.

a Biodistribution at 6, 12 and 24 h post-injection of LNPs (30 μg pDNA (85% Luc+15% Cy5-labeled p1216) per mouse. i.v. n = 2, 36 female BALB/c mice (6–8 weeks) in total). b FACS was used to quantify the percentage of Cy5+ hepatocytes in the liver at 6 and 12 h post-injection. (n = 2) c, d Ex vivo quantitative measurement and luminescence imaging of the liver of BALB/c mice at 24 h post-administration (25 μg Luc pDNA per mouse, intrahepatic injection, n = 2). e–g In vitro transfection and cellular uptake of selected formulations on primary hepatocytes. FACS was used to quantify the percentages of e Cy5+ cells and f GFP+ cells within primary hepatocytes isolated from the liver (1 μg/mL pDNA (75% GFP + 25% Cy5-labeled p1216)). (n = 3) Data are presented as mean values + /− SEM. g Representative FACS data for LNPs pre-incubated with mouse serum for 0.5 h at an LNP/serum volume ratio of 2:1 before dosing. h Quantitative Cellomics high-content analysis for endosomal escape mediated by LNPs in vitro. Average number of Gal8 spots per cell (B16-Gal8-GFP) at 12 h post-treatment as an indication of endosomal escape level (1 μg/mL pDNA) (n = 3). Data are presented as mean values + /− SEM. Data were analyzed using one-way ANOVA and Tukey’s multiple comparisons test (two-sided) for Fig. 6b, c, e, f, h. Statistical P-values: No significance: NS; ^*P < 0.05, ^**P < 0.01, ^***P < 0.001, ^****P < 0.0001. Source data are provided as a Source Data file.