Fig. 3: Experimental and theoretical evaluation of 200-NB activation and its interaction with cells. | Nature Communications

Fig. 3: Experimental and theoretical evaluation of 200-NB activation and its interaction with cells.

From: Light triggered nanoscale biolistics for efficient intracellular delivery of functional macromolecules in mammalian cells

Fig. 3

a Confocal images showing nanoprojectile penetration (indicated with arrow heads) into the cell’s cytoplasm after activation of 200-NBs. The cytoplasm was stained with CellTracker Deep Red. Different vertical sections are shown as well along the y (i and ii) and x (iii and iv) direction. n = 10 images were recorded from two samples. b Flow cytometric quantification of nanoprojectile presence in HeLa cells after 200-NB activation (1.3 × 108 NBs/mL). Samples were irradiated with a laser pulse fluence at 0.5×, 1×, and 1.5× the VB threshold. Controls include 200-NB incubation (1.3 × 108 NBs/mL) without irradiation and the simultaneous incubation of free beads 1% (v/v) and IONPs (1.3 × 108 NPs/mL) with laser irradiation. The percentage of positive cells was quantified by flow cytometry (n = 3 biologically independent samples, data presented as mean ± SD). c Quantification of iron content in HeLa cells after 200-NB activation. (i): Schematic overview of the experimental procedure to determine the iron content in cells (bulk analysis) by ICP-MS. (ii): The iron concentration was measured in untreated cells (negative control), cells incubated with NBs without washing (positive control), and cells treated by activated 200-NBs and subsequent washing with PBS. The results show the mass of Fe normalized per μg of protein as determined by a BSA protein quantification assay (n = 3 biologically independent samples, data presented as mean ± SD, one-way ANOVA, ****P < 0.0001). d Laser-triggered NB-mediated delivery of propidium iodide in HeLa cells. With light-triggered nanobombs PI (yellow) could be successfully delivered into most cells, which was not the case in a control experiment where IONP and nanospheres were added as an uncoupled mixture to the cells. For each condition, n = 10 images were recorded from two samples. The green fluorescent nanospheres are found inside the cells, leading to pore formation in the cell membrane and the concomitant influx of PI that was added to the cell medium. Cell nuclei stained by Hoechst are visible in blue.

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