Imaging intracellular fluorescent proteins at nanometer resolution.
Eric Betzig,George H. Patterson,Rachid Sougrat,O. Wolf Lindwasser,Scott G. Olenych,Juan S. Bonifacino,Michael W. Davidson,Jennifer Lippincott-Schwartz,Harald F. Hess +8 more
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TLDR
This work introduced a method for optically imaging intracellular proteins at nanometer spatial resolution and used this method to image specific target proteins in thin sections of lysosomes and mitochondria and in fixed whole cells to image retroviral protein Gag at the plasma membrane.Abstract:
We introduce a method for optically imaging intracellular proteins at nanometer spatial resolution. Numerous sparse subsets of photoactivatable fluorescent protein molecules were activated, localized (to approximately 2 to 25 nanometers), and then bleached. The aggregate position information from all subsets was then assembled into a superresolution image. We used this method--termed photoactivated localization microscopy--to image specific target proteins in thin sections of lysosomes and mitochondria; in fixed whole cells, we imaged vinculin at focal adhesions, actin within a lamellipodium, and the distribution of the retroviral protein Gag at the plasma membrane.read more
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Using conventional fluorescent markers for far-field fluorescence localization nanoscopy allows resolution in the 10-nm range
Paul Lemmer,Manuel Gunkel,Yanina Weiland,Patrick Müller,David Baddeley,Rainer Kaufmann,Alexander Urich,Heinz Eipel,Roman Amberger,Michael Hausmann,Christoph Cremer +10 more
TL;DR: A novel technique of far‐field localization nanoscopy combining spectral precision distance microscopy with widely used fluorochromes, based on excitation intensity dependent reversible photobleaching of the molecules used combined with fast time sequential imaging under appropriate focusing conditions for laser optical precision localization and image reconstruction with highly enhanced optical resolution in intact cells.
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Non-contact mechanical and chemical analysis of single living cells by microspectroscopic techniques.
Sara Mattana,Maurizio Mattarelli,Lorena Urbanelli,Krizia Sagini,Carla Emiliani,Mauro Dalla Serra,Daniele Fioretto,Silvia Caponi +7 more
TL;DR: In a proof-of-principle experiment, the ability of this spectroscopic technique to characterize subcellular compartments and distinguish cell status was successfully tested and the results strongly support the future application of this technique for fundamental issues in the biomedical field.
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Toward Label-Free Super-Resolution Microscopy
TL;DR: In this paper, the authors combine concepts from Stimulated emission depletion (STED) microscopy and femtosecond Stimulated Raman Spectroscopy (FSRS) for imaging below the diffraction limit.
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A practical guide to optimization in X10 expansion microscopy
TL;DR: This protocol describes an approach for super-resolution imaging by 10× physical expansion of the samples (X10 microscopy) and provides guidelines for determining the expansion and distortion factors.
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Adaptive optics correction of specimen-induced aberrations in single-molecule switching microscopy
TL;DR: In this paper, Adaptive Optics for Single Molecule Switching (SMS) microscopy was proposed to enable the feedback correction of specimen-induced aberrations in SMS microscopy.
References
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Surpassing the lateral resolution limit by a factor of two using structured illumination microscopy.
TL;DR: Lateral resolution that exceeds the classical diffraction limit by a factor of two is achieved by using spatially structured illumination in a wide‐field fluorescence microscope with strikingly increased clarity compared to both conventional and confocal microscopes.
Journal ArticleDOI
Precise nanometer localization analysis for individual fluorescent probes
TL;DR: A localization algorithm motivated from least-squares fitting theory is constructed and tested both on image stacks of 30-nm fluorescent beads and on computer-generated images (Monte Carlo simulations), and results show good agreement with the derived precision equation.
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Nonlinear structured-illumination microscopy: Wide-field fluorescence imaging with theoretically unlimited resolution
TL;DR: Experimental results show that a 2D point resolution of <50 nm is possible on sufficiently bright and photostable samples, and a recently proposed method in which the nonlinearity arises from saturation of the excited state is experimentally demonstrated.
Journal ArticleDOI
Myosin V Walks Hand-Over-Hand: Single Fluorophore Imaging with 1.5-nm Localization
Ahmet Yildiz,Joseph N. Forkey,Sean A. McKinney,Taekjip Ha,Taekjip Ha,Yale E. Goldman,Paul R. Selvin,Paul R. Selvin +7 more
TL;DR: The results strongly support a hand-over-hand model of motility, not an inchworm model, which moves processively on actin.
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