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Nat Methods. 2013 Nov;10(11):1122-6. doi: 10.1038/nmeth.2687. Epub 2013 Oct 6.

Instant super-resolution imaging in live cells and embryos via analog image processing.

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1
Section on High Resolution Optical Imaging, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, Maryland, USA.

Abstract

Existing super-resolution fluorescence microscopes compromise acquisition speed to provide subdiffractive sample information. We report an analog implementation of structured illumination microscopy that enables three-dimensional (3D) super-resolution imaging with a lateral resolution of 145 nm and an axial resolution of 350 nm at acquisition speeds up to 100 Hz. By using optical instead of digital image-processing operations, we removed the need to capture, store and combine multiple camera exposures, increasing data acquisition rates 10- to 100-fold over other super-resolution microscopes and acquiring and displaying super-resolution images in real time. Low excitation intensities allow imaging over hundreds of 2D sections, and combined physical and computational sectioning allow similar depth penetration to spinning-disk confocal microscopy. We demonstrate the capability of our system by imaging fine, rapidly moving structures including motor-driven organelles in human lung fibroblasts and the cytoskeleton of flowing blood cells within developing zebrafish embryos.

PMID:
24097271
PMCID:
PMC3898876
DOI:
10.1038/nmeth.2687
[Indexed for MEDLINE]
Free PMC Article
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