Imaging the nanoscale intracellular structures formed by nucleic acids, such as chromatin, in non-perturbed, structurally and dynamically complex cellular systems, will help improve our understanding of biological processes and open the next frontier for biological discovery. Current optical super-resolution fluorescence techniques require exogenous labels that may disrupt cell function and alter the subdiffractional macromolecular structures they are used to visualize. As a means for label-free optical super-resolution imaging, we examined the discovery of stochastic fluorescence switching of unmodified nucleic acids under visible light illumination. Utilizing this phenomenon and a single-molecule photon localization approach we generated subdiffraction-resolution images down to ~20nm using intrinsic fluorescence from nucleic acids. Specifically, the nanoscale organization of interphase nuclei and mitotic chromosomes were imaged. Using such a method for visualization, we performed a quantitative analysis of the DNA occupancy level and a subdiffractional analysis of the chromosomal organization. These experiments demonstrate a new method for visualizing the nanoscopic features of macromolecular structures composed of nucleic acids without the need for exogenous labels.
Biqin Dong, Luay Matthew Almassalha, Yolanda Stypula-Cyrus, Ben E. Urban, John E. Chandler, The-Quyen Nguyen, Cheng Sun, Hao F. Zhang, and Vadim Backman, "Nanoscopic imaging of chromatin topology utilizing intrinsic fluorescence from unmodified nucleic acids (Conference Presentation)," Proc. SPIE 10071, Single Molecule Spectroscopy and Superresolution Imaging X, 1007109 (Presented at SPIE BiOS: January 28, 2017; Published: 24 April 2017); https://doi.org/10.1117/12.2254900.5380600069001.
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