Figure 1.
Prussian blue staining results of cancer cells internalizing magnetic nanoparticles (MNP) in a time dependent manner.
Control corresponds to the cells (HepG2, SKHep-1, HeLa) that were not treated with MNPs and maintained for 24 h in parallel to the treated groups. Insets: the enlarged images of cells. Blue: MNPs; Red: nucleus. The scale bar in each image represents 40 µm.
Figure 2.
Magnetophoresis and living cell imaging.
(a) Schematic representation of cell magnetophoresis. Fmag and Fvis indicate the magnetic force and the viscous force. (b) (Top) Consecutive magnetophoresis images of two cells move at constant speed (Bottom) Fluorescence image, optical microscope image and the merged image of the two cells. Green, live cells expressed green fluorescent protein (GFP); Red, nucleic acid stained by propidium iodide (PI). The scale bar represents 20 µm. (c) Size distributions of distinctive cells (SKHep-1, HepG2 and HeLa). The red lines are the standard normal distribution. N = 200 cells.
Figure 3.
Quantitative analysis of magnetic nanoparticle (MNP) monitored as a function of the incubation time.
(a) The distributions of magnetophoretic velocity of cells incubated with magnetic nanoparticles for 8, 12, 24 h. N = 1000. (b) The cell loading with MNPs after incubating for 8, 12 and 24 h. Dot lines represent the exponential fit of the data. Bars represent the standard deviation of 500 different cells.
Figure 4.
Cytotoxicity effects of single cell and bulk cell populations.
(a) Cytotoxicity effects were analyzed by MTT. The data represents as the mean ±SD, and acquired from 3 independent tests. (b) The cytotoxicity effects of HeLa cells are shown by the overlay fluorescence images of GFP and PI as the MNP incubation time increased from 8 to 24 h. Green, live cells expressed green fluorescent protein (GFP); Red, dead cells stained by PI. The scale bar represents 40 µm. (c) Overlay fluorescence images of DAPI/PI stain distinguishing mechanisms of cell death (SKHep-1, HepG2 and HeLa) after treating with MNPs for 24 h. Control: cells were not treated with MNPs for 24 h in parallel to the treated groups. Apoptotic nuclei stained with DAPI showed intense blue fluorescence corresponding to chromatin condensation (indicated by arrow heads) and fragmentation (indicated by arrows). Defective cell membrane allowed PI to enter cytoplasm indicating that cells underwent late apoptosis or necrosis. Possible necrotic cell that correspond to those without clear nuclear fragment but stained with intense PI were circled. (d) Time course experiment of the DAPI/PI staining for the three kinds of cells.
Table 1.
The mean percentage of normal, apoptotic and necrotic cell stained with DAPI/PI.
Figure 5.
The correlation between the number of internalized MNPs and cell viability.
The three data points for each cell type represent the three incubation times (8, 12, 24 h) with MNPs. The ratio of survival to total cell number was regarded as viability (PI-positive cells with cell necrosis or late apoptotic nuclei were regarded as dead cells). The error bars represent the standard deviations (SD) obtained from three independent replicates. The correlation between the mean MNPs number obtained from magnetophoresis and the viability of the cells was determined by linear regression.
Figure 6.
The variability of the number of intracellular MNPs obtained from live (green) and dead (red) cell.
(a)∼(c) are box plots of HeLa, HepG2 and SKHep-1, respectively. For each cell type, 500 cells that were treated with MNPs were analyzed at different co-culture times (8, 12, 24 h) to obtain the uptaken MNP number and viability of an individual cell. (d) is the data collection of all 1500 cells with different co-culture times (8, 12, 24 h) for each cell type. The dead/live cells were discriminated by the fluorescence images in the magnetophoresis experiment. The box represents the quartile distribution (25–75%) range. The median is indicated by a horizontal line and the mean is indicated by an open square inside the box. The whiskers mark 5–95% range of the distribution, and the outlier samples are indicated outside of this range. The scatter plots indicate the uptaken MNP number obtained from individual cell. Statistical significance between dead and live cells was determined using a student's t-test. The degree of significance was given as *: p<0.05; **: p<0.01; ***: p<0.001.