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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Journal ArticleDOI
Single-molecule imaging of UvrA and UvrB recruitment to DNA lesions in living Escherichia coli
Mathew Stracy,Marcin Jaciuk,Stephan Uphoff,Achillefs N. Kapanidis,Marcin Nowotny,David J. Sherratt,Pawel Zawadzki +6 more
TL;DR: Single-molecule fluorescence imaging is used to provide a comprehensive characterization of the lesion search, recognition and verification process in living cells and shows that NER initiation involves a two-step mechanism in which UvrA scans the genome and locates DNA damage independently of UvrB.
Journal ArticleDOI
Challenges in quantitative single molecule localization microscopy
TL;DR: This review article focuses specifically on Photo‐Activated Localization Microscopy (PALM), due to its advantages in labeling specificity and the relatively low overcounting caused by photoblinking when photo‐activable fluorescent proteins are used as labels.
Journal ArticleDOI
TCRs are randomly distributed on the plasma membrane of resting antigen-experienced T cells.
Benedikt K. Rossboth,Andreas M. Arnold,Haisen Ta,René Platzer,Florian Kellner,Johannes B. Huppa,Mario Brameshuber,Florian Baumgart,Gerhard J. Schütz +8 more
TL;DR: It is found here that apparent TCR nanoclustering could be attributed to overcounting artifacts inherent to single-molecule-localization microscopy, and the distribution of TCRs on the plasma membrane is optimized for fast recognition of antigen in the first phase of T cell activation.
Journal ArticleDOI
The 2018 correlative microscopy techniques roadmap.
Toshio Ando,Satya Prathyusha Bhamidimarri,Niklas Brending,Huw Colin-York,Lucy M. Collinson,Niels de Jonge,Niels de Jonge,P. J. de Pablo,Elke Debroye,Christian Eggeling,Christian Eggeling,Christian Eggeling,Christian Franck,Marco Fritzsche,Hans C. Gerritsen,Ben N G Giepmans,Kay Grünewald,Johan Hofkens,Jacob P. Hoogenboom,Kris P. F. Janssen,Rainer Kaufman,Judith Klumpermann,Nyoman D. Kurniawan,Jana Kusch,Nalan Liv,Viha Parekh,Diana B. Peckys,Florian Rehfeldt,David C. Reutens,Maarten B. J. Roeffaers,Tim Salditt,Iwan A. T. Schaap,Ulrich S. Schwarz,Paul Verkade,Michael W. Vogel,Richard Wagner,Mathias Winterhalter,Haifeng Yuan,Giovanni Zifarelli +38 more
TL;DR: A special roadmap on the correlative microscopy techniques is presented, giving a comprehensive overview from various leading scientists in this field, via a collection of multiple short viewpoints.
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A Nebulin Ruler Does Not Dictate Thin Filament Lengths
TL;DR: The results suggest the existence of a mechanism whereby nebulin specifies the minimum thin filament length and sarcomere length regulates and coordinates pointed-end dynamics to maintain the relative overlap of the thin and thick filaments during myofibril assembly.
References
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疟原虫var基因转换速率变化导致抗原变异[英]/Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A
TL;DR: PfPMP1)与感染红细胞、树突状组胞以及胎盘的单个或多个受体作用,在黏附及免疫逃避中起关键的作�ly.
Journal ArticleDOI
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.
Journal ArticleDOI
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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