Three-Dimensional Super-Resolution Imaging by Stochastic Optical Reconstruction Microscopy
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Cites methods from "Three-Dimensional Super-Resolution ..."
...The “Z Calibration” button assumes the use of an astigmatic lens introduced in the imaging path to measure Z position by the shape of the fit Gaussian [11]....
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...Smaller probes include primary antibodies for direct immunofluorescence (~10–15 nm in size) (52), antibody Fab fragments (~6 nm in size), and FPs (~3–4 nm in size)....
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...This method has achieved a lateral localization precision of ~25 nm (FWHM) and an axial localization precision of ~50 nm (FWHM) over the depth of several hundred nanometers in z with photoswitchable cyanine dyes (52)....
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...The cytoskeleton of mammalian cells, especially microtubules (Figure 5a) (29, 44, 46, 52), is the most commonly used benchmark structure for super-resolution imaging....
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...These images have shown the ability of super-resolution fluorescence microscopy to separate nearby filaments and overlapped and unresolvable in conventional fluorescence images (46, 52), as well as to reveal the 3D organization of the cytoskeleton in a mammalian cell (52)....
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...The spatial organization of microtubules in cells, the semispherical shell structure of clathrin-coated pits, with a diameter of about ~150 nm, and the outer membrane structure of mitochondria has been resolved using this technique (52,53) For thick sample imaging, spherical abberations caused by the refractive index mismatch between the sample and the imaging optics also need to be minimized or corrected (53)....
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References
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