4.6 Article

Method for nanoparticles uptake evaluation based on double labeled fluorescent cells scanned in enhanced darkfield microscopy

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BIOMEDICAL OPTICS EXPRESS
卷 14, 期 6, 页码 2796-2810

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Optica Publishing Group
DOI: 10.1364/BOE.490136

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We propose a method that combines imaging tools and computational tools to locate and detect unlabeled nanoparticles (NPs) in cells and evaluate their internal traffic. The method utilizes a dark field CytoViva optical system and combines 3D reconstructions of double fluorescently labeled cells with hyperspectral images. MATLAB scripts are used to process the images and localize NPs in different cell regions. The method provides important parameters to assess uptake efficiency and shows consistent results with biochemical analyses.
We present a method that integrates the standard imaging tools for locating and detecting unlabeled nanoparticles (NPs) with computational tools for partitioning cell volumes and NPs counting within specified regions to evaluate their internal traffic. The method uses enhanced dark field CytoViva optical system and combines 3D reconstructions of double fluorescently labeled cells with hyperspectral images. The method allows the partitioning of each cell image into four regions: nucleus, cytoplasm, and two neighboring shells, as well as investigations across thin layers adjacent to the plasma membrane. MATLAB scripts were developed to process the images and to localize NPs in each region. Specific parameters were computed to assess the uptake efficiency: regional densities of NPs, flow densities, relative accumulation indices, and uptake ratios. The results of the method are in line with biochemical analyses. It was shown that a sort of saturation limit for intracellular NPs density is reached at high extracellular NPs concentrations. Higher NPs densities were found in the proximity of the plasma membranes. A decrease of the cell viability with increasing extracellular NPs concentration was observed and explained the negative correlation of the cell eccentricity with NPs number.& COPY; 2023 Optica Publishing Group under the terms of the Optica Open Access Publishing Agreement

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