Fig. 2.

Physicochemical properties and HCC cell transfection of scAb-EGFR-targeted nanomedicine. a Determination of plasmid DNA (pDNA) complexation at various nitrogen/phosphorus (N/P) ratios by gel retardation assay. Full complexation with single-chain antibody (scAb)-EGFR-PEG-g-PEI-SPION prevents DNA migration into the gel. b Particle size and zeta potential of scAb-EGFR-PEG-g-PEI-SPION/pDNA at various N/P ratios. Data were represented as means ± S.D. Error bars represent the S.D. values (n = 3). c Transmission electron microscopy (TEM) images of nanomedicine scAb-EGFR-PEG-g-PEI-SPION/pDNA. The gray halo around the dark SPION cluster indicates a polymeric shell. Scale bar, 100 nm. d T2-weighted magnetic resonance imaging (MRI) examination of different groups of Hep3B cells after 1 h of incubation at Fe concentrations of 0, 5, 10, 20, 40, and 80 μg/mL. e Left four panels, confocal laser scanning microscopic (CLSM) imaging of different groups of Hep3B cells incubated for 0.25, 1, 2, 4 h, respectively, at Fe concentration of 20 μg/mL. Scale bar, 10 μm. (Blue, DAPI-indicating nuclei; red, POPO-3 indicating pDNA; green, Oregon Green 488-indicating delivery agent; Overlapping of red and green fluorescence to generate a yellow one, see also Supplementary Fig. 1d). Right panel, Prussian blue staining images of different groups of Hep3B cells incubated at the same conditions as in CLSM studies. Scale bar, 50 μm. (Blue, indicating Fe aggregation; red, indicating nuclei). f Protein expression levels of PBOV1 in Hep3B, SMMC-7721, and BEL7402 cells relative to β-actin as control determined by western blotting assay