期刊
NATURE PROTOCOLS
卷 16, 期 1, 页码 383-404出版社
NATURE RESEARCH
DOI: 10.1038/s41596-020-00420-1
关键词
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资金
- National Natural Science Foundation of China [91953106, 21904041, 21804088]
- China Postdoctoral Science Foundation [2019M661417]
This method provides a step-by-step correlative imaging approach to investigate plasmonic NPs of different aggregation states in single cells. By using dark-field microscopy, fluorescence microscopy, and scanning electron microscopy, the clustering states of SNAs in a single cell can be efficiently distinguished and tracked in real time within 20 seconds. The entire procedure can be completed in 3-5 days and requires standard imaging and data analysis skills and equipment.
A comprehensive understanding of interactions between nanoparticles (NPs) and biological components is critical to the clinical application of NPs and nanomedicine. Here we provide a step-by-step correlative imaging approach to investigate plasmonic NPs of different aggregation states at the single-cell level. Traceable spherical nucleic acids (SNAs) are fabricated by decorating 50-nm spherical gold NPs with fluorophore-labeled DNA, serving as dually emissive (fluorescent and plasmonic) NPs. The in situ correlative imaging with dark-field microscopy (DFM) and fluorescence microscopy (FM) reveals intracellular distribution of SNAs, whereas DFM combined with scanning electron microscopy (SEM) allows semi-quantification of SNA clustering states in solution. The imaging data are analyzed by ImageJ and a colorimetry-based algorithm written in Python. The clustering states of SNAs in a single cell can be efficiently distinguished within 20 s. This method can be readily installed to monitor real-time endocytosis and cellular distribution of plasmonic NPs of different aggregation states and to quantitatively image targets of interest (e.g., specific DNA, messenger RNA, peptides or proteins) in living cells. The entire procedure can be completed in 3-5 d and requires standard DFM, FM and SEM imaging and data analysis skills and equipment. We report a new protocol for tracking endocytosis and cellular distribution of spherical nucleic acid nanoparticles of different clustering states by correlative imaging with dark-field microscopy, fluorescence microscopy and scanning electron microscopy.
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