Synthesis and in vitro visualization of fluorescein isothiocyanate-labeled chitosan nanoparticles (fitc-cnp) in human kidney cancer cells

The use of fluorescently-labeled nanoparticles has garnered attention in the medical field, due to the ability to track its uptake during cell transfection. In this study, the synthesis and in vitro visualization of fluorescein isothiocyanate (FITC)-labeled chitosan nanoparticles (CNPs) inside human kidney cancer cells (786-O cell line) were described. CNPs were prepared using a modified ionic gelation method to yield spherical nanoparticles, and fluorescent labeling of CNP was performed using N-hydroxy-succinimidyl (NHS)-FITC. Particle size distribution and polydispersity index of nanoparticles were analyzed using dynamic light scattering. Surface morphology was analyzed using FE-SEM. For transfection, 786-O human kidney cancer cells were established and subsequently treated with both CNP and FITC-CNP, visualized at three different time points using fluorescence microscopy. It was found that 600 μl of 0.5 mg/ml chitosan with 200 μl of 0.7 mg/ml TPP gave the optimum particle size and PDI value at 71.7+0.4 nm and 0.14+0.01, respectively. FE-SEM data obtained was consistent with DLS results at optimum parameters. Visualization of cell treatment showed that at 30 mins, all FITC-CNPs were seen as small green dots with weak fluorescence residing outside of the cells at close proximity to the cell membrane. Most of the nanoparticles internalized into the cells at 6h were seen to be in the cytoplasm or surrounding the nucleus. As time progresses to 24h, stronger fluorescence was observed with all FITC-CNPs accumulating inside cells with smearing observed, probably due to FITC detachment from CNPs. These results implied that the synthesized FITC-labeled CNPs have the potential for use as carrier system for enhanced drug delivery into cells.

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