Determining the structure and composition of macromolecular assemblies is a major challenge in biology. Here we describe ultrastructure expansion microscopy (U-ExM), an extension of expansion microscopy that allows the visualization of preserved ultrastructures by optical microscopy. This method allows for near-native expansion of diverse structures in vitro and in cells; when combined with super-resolution microscopy, it unveiled details of ultrastructural organization, such as centriolar chirality, that could otherwise be observed only by electron microscopy.

Abberior Instruments GmbH Göttingen Germany

Department of Biotechnology and Biophysics Biocenter University of Würzburg Würzburg Germany

Department of Cell Biology Sciences 3 University of Geneva Geneva Switzerland

Ecole Polytechnique Fédérale de Lausanne Biomedical Imaging Group Lausanne Switzerland

ICube CNRS University of Strasbourg Illkirch France

Laboratory of Adaptative Immunity Institute of Molecular Genetics Academy of Sciences of the Czech Republic Prague Czech Republic

Massachusetts Institute of Technology Cambridge MA USA

Signal Processing Core of Center for Biomedical Imaging EPFL Lausanne Switzerland

Zobrazit více v PubMed

Koster AJ, Klumperman J. Electron microscopy in cell biology: integrating structure and function. Nat Rev Mol Cell Biol. 2003;Suppl:SS6–10. PubMed

Sahl SJ, Hell SW, Jakobs S. Fluorescence nanoscopy in cell biology. Nat Rev Mol Cell Biol. 2017;18:685–701. PubMed

Chen F, Tillberg PW, Boyden ES. Optical imaging. Expansion microscopy. Science. 2015;347:543–8. PubMed PMC

Chozinski TJ, et al. Expansion microscopy with conventional antibodies and fluorescent proteins. Nat Methods. 2016;13:485–488. PubMed PMC

Tillberg PW, et al. Protein-retention expansion microscopy of cells and tissues labeled using standard fluorescent proteins and antibodies. Nat Biotechnol. 2016;34:987–992. PubMed PMC

Ku T, et al. Multiplexed and scalable super-resolution imaging of three-dimensional protein localization in size-adjustable tissues. Nat Biotechnol. 2016;34:973–981. PubMed PMC

Hamel V, et al. Identification of Chlamydomonas Central Core Centriolar Proteins Reveals a Role for Human WDR90 in Ciliogenesis. Curr Biol. 2017;27:2486–2498.e6. PubMed PMC

Klena N, et al. Isolation and Fluorescence Imaging for Single-particle Reconstruction of Chlamydomonas Centrioles. JoVE. 2018:e58109. doi: 10.3791/58109. PubMed DOI PMC

Heilemann M, et al. Subdiffraction-Resolution Fluorescence Imaging with Conventional Fluorescent Probes **. 2008:6172–6176. doi: 10.1002/anie.200802376. PubMed DOI

Fortun D, et al. Reconstruction from Multiple Particles for 3D Isotropic Resolution in Fluorescence Microscopy. IEEE Trans Med Imaging. 2018:1–1. doi: 10.1109/TMI.2018.2795464. PubMed DOI

Heine J, et al. Adaptive-illumination STED nanoscopy. Proc Natl Acad Sci. 2017 doi: 10.1073/pnas.1708304114. 201708304. PubMed DOI PMC

Pigino G, et al. Cryoelectron tomography of radial spokes in cilia and flagella. J Cell Biol. 2011;195:673–87. PubMed PMC

Lechtreck KF, Geimer S. Distribution of polyglutamylated tubulin in the flagellar apparatus of green flagellates. Cell Motil Cytoskeleton. 2000;47:219–235. PubMed

Kubo T, Yanagisawa H aki, Yagi T, Hirono M, Kamiya R. Tubulin Polyglutamylation Regulates Axonemal Motility by Modulating Activities of Inner-Arm Dyneins. Curr Biol. 2010;20:441–445. PubMed

Kubo T, Oda T. Electrostatic interaction between polyglutamylated tubulin and the nexin–dynein regulatory complex regulates flagellar motility. Mol Biol Cell. 2017;28:2260–2266. PubMed PMC

Suryavanshi S, et al. Tubulin Glutamylation Regulates Ciliary Motility by Altering Inner Dynein Arm Activity. Curr Biol. 2010;20:435–440. PubMed PMC

Gao M, et al. Expansion Stimulated Emission Depletion Microscopy (ExSTED) ACS Nano. 2018;12:4178–4185. PubMed

Halpern AR, Alas GCM, Chozinski TJ, Paredez AR, Vaughan JC. Hybrid Structured Illumination Expansion Microscopy Reveals Microbial Cytoskeleton Organization. ACS Nano. 2017;11:12677–12686. PubMed PMC

Yang TT, et al. Super-resolution architecture of mammalian centriole distal appendages reveals distinct blade and matrix functional components. Nat Commun. 2018;9:1–11. PubMed PMC

Borlinghaus RT, Kappel C. HyVolution—the smart path to confocal super-resolution. Nat Methods. 2016;13:i–iii.

de Luca GMR, et al. Configurations of the Re-scan Confocal Microscope (RCM) for biomedical applications. J Microsc. 2017;266:166–177. PubMed

Göttfert F, et al. Strong signal increase in STED fluorescence microscopy by imaging regions of subdiffraction extent. Proc Natl Acad Sci. 2017;114:2125–2130. PubMed PMC

Ovesný M, Křížek P, Borkovec J, Švindrych Z, Hagen GM. ThunderSTORM: A comprehensive ImageJ plug-in for PALM and STORM data analysis and super-resolution imaging. Bioinformatics. 2014;30:2389–2390. PubMed PMC

Wolter S, et al. rapidSTORM : accurate, fast open-source software for localization microscopy orcae : online resource for community annotation of eukaryotes. Nat Methods. 2012;9:1040–1. PubMed

Thevenaz P, Ruttimann UE, Unser M. A pyramid approach to subpixel registration based on intensity. IEEE Trans Image Process. 1998;7:27–41. PubMed

Unser M, Soubies E, Soulez F, McCann M, Donati L. GlobalBioIm: A Unifying Computational Framework for Solving Inverse Problems. Imaging Appl Opt 2017 (3D, AIO, COSI, IS, MATH, pcAOP) 2017;2017 CTu1B.1.

Schindelin J, et al. Fiji: An open-source platform for biological-image analysis. Nat Methods. 2012;9:676–682. PubMed PMC

Schneider Ca, Rasband WS, Eliceiri KW. NIH Image to ImageJ: 25 years of image analysis. Nat Methods. 2012;9:671–675. PubMed PMC

Royer LA, et al. ClearVolume: Open-source live 3D visualization for light-sheet microscopy. Nat Methods. 2015;12:480–481. PubMed

Visualisation of Euglena gracilis organelles and cytoskeleton using expansion microscopy

Detailed characterisation of the trypanosome nuclear pore architecture reveals conserved asymmetrical functional hubs that drive mRNA export

Detection of TurboID fusion proteins by fluorescent streptavidin outcompetes antibody signals and visualises targets not accessible to antibodies

ULK4 and Fused/STK36 interact to mediate assembly of a motile flagellum

Imaging plant cells and organs with light-sheet and super-resolution microscopy

Expansion microscopy facilitates quantitative super-resolution studies of cytoskeletal structures in kinetoplastid parasites

Prospects and limitations of expansion microscopy in chromatin ultrastructure determination

The BBSome assembly is spatially controlled by BBS1 and BBS4 in human cells