Abstract
Precise localization of nanoparticles within a cell is crucial to the understanding of cell-particle interactions and has broad applications in nanomedicine. Here, we report a proof-of-principle experiment for imaging individual functionalized nanoparticles within a mammalian cell by correlative microscopy. Using a chemically-fixed HeLa cell labeled with fluorescent core-shell nanoparticles as a model system, we implemented a graphene-oxide layer as a substrate to significantly reduce background scattering. We identified cellular features of interest by fluorescence microscopy, followed by scanning transmission X-ray tomography to localize the particles in 3D, and ptychographic coherent diffractive imaging of the fine features in the region at high resolution. By tuning the X-ray energy to the Fe L-edge, we demonstrated sensitive detection of nanoparticles composed of a 22 nm magnetic Fe3O4 core encased by a 25-nm-thick fluorescent silica (SiO2) shell. These fluorescent core-shell nanoparticles act as landmarks and offer clarity in a cellular context. Our correlative microscopy results confirmed a subset of particles to be fully internalized, and high-contrast ptychographic images showed two oxidation states of individual nanoparticles with a resolution of ~16.5 nm. The ability to precisely localize individual fluorescent nanoparticles within mammalian cells will expand our understanding of the structure/function relationships for functionalized nanoparticles.
Highlights
Functionalized nanoparticles are used in a broad array of nanomedicine applications for their utility as labels and drug delivery systems[1,2,3,4]
We demonstrate a correlative ptychographic approach for high-resolution imaging of functionalized nanoparticles internalized within an un-sectioned mammalian cell
Graphene and graphene-oxide films have previously been shown to be an excellent support for high-resolution electron microscopy studies of cells and macromolecules, both frozen-hydrated[45] and in-situ[46,47,48]
Summary
Nano-labeling of HeLa cells with fluorescent nanoparticles on a graphene-oxide substrate. We achieved high-resolution imaging of the leading edge of a HeLa cell treated with fluorescent core-shell Fe3O4-SiO2 nanoparticles by correlative microscopy. To reveal the local distribution of fluorescent Fe3O4-SiO2 nanolabels in or near the HeLa cell, we performed 2D ptychographic CDI on regions of interest identified from the 3D reconstruction using STXM tomography (Fig. 3a–c). The ability to perform correlative cellular imaging and localize individual nanoparticles inside intact, un-sectioned mammalian cells through a combination of fluorescent microscopy, STXM tomography and ptychography will yield a more comprehensive understanding of the cell as a complex biological system, and find applications in quantifying cell-particle interactions in nanomedicine
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