Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 109, Issue 35, Pages 13978-13983Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1201882109
Keywords
nanoscopy; diffraction limit; photoswitchable dye; stochastic optical reconstruction microscopy; photoactivation localization microscopy
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Funding
- US National Institutes of Health
- Howard Hughes Medical Institute
- National Natural Science Foundation of China
- National Basic Research Program of China
- Mary Fieser fellowship
- Burroughs-Wellcome Career Award at the Scientific Interface
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Imaging membranes in live cells with nanometer-scale resolution promises to reveal ultrastructural dynamics of organelles that are essential for cellular functions. In this work, we identified photoswitchable membrane probes and obtained super-resolution fluorescence images of cellular membranes. We demonstrated the photoswitching capabilities of eight commonly used membrane probes, each specific to the plasma membrane, mitochondria, the endoplasmic recticulum (ER) or lysosomes. These small-molecule probes readily label live cells with high probe densities. Using these probes, we achieved dynamic imaging of specific membrane structures in living cells with 30-60 nm spatial resolution at temporal resolutions down to 1-2 s. Moreover, by using spectrally distinguishable probes, we obtained two-color super-resolution images of mitochondria and the ER. We observed previously obscured details of morphological dynamics of mitochondrial fusion/fission and ER remodeling, as well as heterogeneous membrane diffusivity on neuronal processes.
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