LNP uptake and eGFP expression
in Huh7 cells exposed to LNPs freshly
diluted in 10% Lipo-D or 10% FBS. (A) Representative confocal microscopy
images of cells treated as mentioned above, including negative control
(LNP-untreated cells). The images were acquired and treated with identical
settings, and intensities are thus comparable. All scale bars are
50 μm. (B–D) Cell uptake (Cy5 signal, B), protein expression
(eGFP signal, C), and endosomal escape efficiency (Cy5/eGFP signal,
D), measured by flow cytometry (N = 3; error bars
represent standard deviation of three independent biological replicates).
The cells were incubated with LNPs at a mRNA concentration of 0.625
μg/mL for 2 h prior to analysis to allow for detectable levels
of uptake and protein expression. Means comparison for data in B–D
was performed by an unpaired Student’s t test, N = 3; * and *** denote statistically significant differences
in mean at the p < 0.05 and p < 0.0001 levels, respectively. The p value for
means comparison for the data in (B) was 0.037.