Figure 1.
Schematic illustration of the MagNP-based method of multiple-gene delivery into porcine kidney cells.
After complexation of plasmids containing the genes encoding GFP and DsRed to the MagNPs, the MagNP-DNA complexes were added to the cells and a magnetic force (Fmag) was applied to promote gene delivery.
Figure 2.
SEM images of PEI-coated Fe3O4 nanoparticles.
(a,b) The high concentration (50 µg/ml) sample of the PEI-coated MagNPs showing slight aggregation of the nanoparticles. The inset shows a single spherical nanoparticle. (b) Higher magnification of the low concentration (10 µg/ml) sample of the PEI-coated MagNPs.
Figure 3.
Agarose gel electrophoresis of MagNP-DNA complexes with different DNA:MagNP weight ratios.
(a) Migration of MagNP-DNAGFP complexes with DNA:MagNP weight ratios of 20∶1, 10∶1, 5∶1, 1∶1, 0.2∶1, and 0.1∶1. (b) Corresponding 3D projection of Figure 3a. (c) Migration of MagNP-DNADsRed complexes with the same DNA:MagNP weight ratios. (d) Corresponding 3D projection of Figure 3c. Pure DNA plasmids were used in each case as controls.
Figure 4.
AFM images of MagNPs and MagNP-DNA complexes.
(a–c) AFM images of MagNPs. (a) The red arrowheads indicate the diameter of the MagNPs and the inset shows the corresponding phase image. (b) The topographic distance profile corresponding to the region between the red arrowheads in Figure 4a. (c) 3D rendering of the individual MagNPs shown in Figure 4a. (d–f) AFM images of MagNP-DNA complexes. (d) MagNPs are bound to the stretched DNA strands. The local and representative DNA strands are shown in the inset. The red arrowheads indicate the height of the DNA strands. (e) Phase image of MagNP-DNA complexes. (f) 3D rendering of the MagNP-DNA complexes shown in Figure 4d.
Figure 5.
Histograms of the particle size distribution of MagNPs and MagNP-DNA complexes.
(a) MagNPs. (b) MagNP-DNAGFP complexes. (c) MagNP-DNADsRed complexes.
Figure 6.
Fluorescence microscopy images of GFP and DsRed expression in transfected PK-15 cells.
Green or red fluorescence was detected 24-DNAGFP (a–c) or MagNP-DNADsRed (d–f) complexes, respectively. (Scale bars, 100 µm).
Figure 7.
Fluorescence microscopy analyses of the localization of exogenous GFP in transfected PK-15 cells.
PK-15 cells magnofected with the MagNP-DNAGFP complex were stained with a membrane-specific red fluorescent dye (DiI) and a nucleus-specific blue fluorescent dye (DAPI) 24 h after transfection. (a) Merged image showing the red membrane, blue nucleus, and GFP (green) expression. (b) DAPI staining only. (c) DiI staining only. (d) GFP signal only. (Scale bars, 20 µm).
Figure 8.
Fluorescence microscopy analyses of co-expressed GFP and DsRed in transfected PK-15 cells.
PK-15 cells were co-magnofected with the MagNP-DNAGFP and MagNP-DNADsRed complexes and images were collected 24 h after transfection. (a–d) Fluorescence (a–c) and bright field imaging (d) of the cells spread between two glass cover slips. GFP and DsRed fluorescence were detected in the green (500–530 nm) and red (552–617 nm) channels, respectively. (Scale bars, 20 µm).
Figure 9.
Fluorescence microscopy analyses of GFP and DsRed in PK-15 cells co-transfected without a magnetic field.
PK-15 cells were co-transfected with the MagNP-DNAGFP and MagNP-DNADsRed complexes in the absence of an external magnetic field and images were collected 24 h after transfection. (a–d) Fluorescence (a–c) and bright field imaging (d) of the cells spread between two glass cover slips. GFP and DsRed fluorescence were detected in the green (500–530 nm) and red (552–617 nm) channels, respectively. (Scale bars, 50 µm).
Figure 10.
Flow cytometry analysis of the co-expression of GFP and DsRed in transfected PK-15 cells.
Cells expressing GFP, DsRed, and GFP plus DsRed are shown in the lower right (LR), upper left (UL), and upper right (UR) quadrants, respectively. The percentages of cells in each quadrant are shown in the table on the right.