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Cited by 140Advanced Electron Microscopy Techniques and ApplicationsAdvanced Fluorescence Microscopy TechniquesAdvanced Materials Characterization Techniques

Correlated cryo-fluorescence and cryo-electron microscopy with high spatial precision and improved sensitivity

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Schorb, Martin, Briggs, John A.G.·2014·Ultramicroscopy
DOI 10.1016/j.ultramic.2013.10.015PMC PMC5472196
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Abstract

Performing fluorescence microscopy and electron microscopy on the same sample allows fluorescent signals to be used to identify and locate features of interest for subsequent imaging by electron microscopy. To carry out such correlative microscopy on vitrified samples appropriate for structural cryo-electron microscopy it is necessary to perform fluorescence microscopy at liquid-nitrogen temperatures. Here we describe an adaptation of a cryo-light microscopy stage to permit use of high-numerical aperture objectives. This allows high-sensitivity and high-resolution fluorescence microscopy of vitrified samples. We describe and apply a correlative cryo-fluorescence and cryo-electron microscopy workflow together with a fiducial bead-based image correlation procedure. This procedure allows us to locate fluorescent bacteriophages in cryo-electron microscopy images with an accuracy on the order of 50 nm, based on their fluorescent signal. It will allow the user to precisely and unambiguously identify and locate objects and events for subsequent high-resolution structural study, based on fluorescent signals.

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  • The accuracy of the correlation was measured by leave-one-out calculations as described by Schorb and Briggs (2014).

    “Leave-one-out calculations were performed as described by Schorb and Briggs (2014).”

    From StableMARK-decorated microtubules in cells have expanded lattices (2025) · cites this paper as Schorb and Briggs (2014)

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  1. [1]Tsien RY The green fluorescent protein Annu Rev Biochem 1998 67 509 544 9759496
  2. [2]Giepmans BNG Adams SR Ellisman MH Tsien RY The fluorescent toolbox for assessing protein location and function Science 2006 312 217 224 16614209
  3. [3]Betzig E Patterson GH Sougrat R Lindwasser OW Olenych S Bonifacino JS Imaging intracellular fluorescent proteins at nanometer resolution Science 2006 313 1642 1645 16902090
  4. [4]Klar TA Jakobs S Dyba M Egner A Hell SW Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission Proc Natl Acad Sci U S A 2000 97 8206 8210 10899992 PMC26924
  5. [5]Karreman MA Agronskaia AV van Donselaar EG Vocking K Fereidouni F Humbel BM Optimizing immuno-labeling for correlative fluorescence and electron microscopy on a single specimen J Struct Biol 2012 180 382 386 22982545
  6. [6]Bouchet-Marquis C Pagratis M Kirmse R Hoenger A Metallothionein as a clonable high-density marker for cryo-electron microscopy J Struct Biol 2012 177 119 127 22068155 PMC3261350
  7. [7]Risco C Sanmartín-Conesa E Tzeng W-P Frey TK Seybold V de Groot RJ Specific, sensitive, high-resolution detection of protein molecules in eukaryotic cells using metal-tagging transmission electron microscopy Struct Lond Engl 2012 20 759 766 1993 PMC3351761 22579245
  8. [8]Brown E Verkade P The use of markers for correlative light electron microscopy Protoplasma 2010 244 91 97 20524017
  9. [9]van Rijnsoever C Oorschot V Klumperman J Correlative light-electron microscopy (CLEM) combining live-cell imaging and immunolabeling of ultrathin cryosections Nat Methods 2008 5 973 980 18974735
  10. [10]Polishchuk RS Polishchuk EV Marra P Alberti S Buccione R Luini A Correlative light-electron microscopy reveals the tubular-saccular ultrastructure of carriers operating between Golgi apparatus and plasma membrane J Cell Biol 2000 148 45 58 10629217 PMC2156208
  11. [11]Spiegelhalter C Tosch V Hentsch D Koch M Kessler P Schwab Y From Dynamic Live Cell Imaging to 3D Ultrastructure: Novel Integrated Methods for High Pressure Freezing and Correlative Light-Electron Microscopy PLoS One 2010 5 PMC2815783 20140253
  12. [12]Müller-Reichert T Srayko M Hyman A O’Toole ET McDonald K Correlative light and electron microscopy of early Caenorhabditis elegans embryos in mitosis Methods Cell Biol 2007 79 101 119 17327153
  13. [13]Kolotuev I Bumbarger DJ Labouesse M Schwab Y Targeted ultramicrotomy: a valuable tool for correlated light and electron microscopy of small model organisms Methods Cell Biol 2012 111 203 222 22857930
  14. [14]McDonald KL A review of high-pressure freezing preparation techniques for correlative light and electron microscopy of the same cells and tissues J Microsc 2009 235 273 281 19754722
  15. [15]Nixon SJ Webb RI Floetenmeyer M Schieber N Lo HP Parton RG A single method for cryofixation and correlative light, electron microscopy and tomography of zebrafish embryos Traffic Cph Den 2009 10 131 136 19054388
  16. [16]Kukulski W Schorb M Welsch S Picco A Kaksonen M Briggs JAG Correlated fluorescence and 3D electron microscopy with high sensitivity and spatial precision J Cell Biol 2011 192 111 119 21200030 PMC3019550
  17. [17]Kukulski W Schorb M Welsch S Picco A Kaksonen M Briggs JAG Precise, correlated fluorescence microscopy and electron tomography of lowicryl sections using fluorescent fiducial markers Methods Cell Biol 2012 111 235 257 22857932
  18. [18]Briegel A Chen S Koster AJ Plitzko JM Schwartz CL Jensen GJ Correlated light and electron cryo-microscopy Methods Enzymol 2010 481 317 341 20887863
  19. [19]Sartori A Gatz R Beck F Rigort A Baumeister W Plitzko JM Correlative microscopy: bridging the gap between fluorescence light microscopy and cryo-electron tomography J Struct Biol 2007 160 135 145 17884579
  20. [20]Rigort A Villa E Bäuerlein FJB Engel BD Plitzko JM Integrative approaches for cellular cryo-electron tomography: correlative imaging and focused ion beam micromachining Methods Cell Biol 2012 111 259 281 22857933
  21. [21]Schwartz CL Sarbash VI Ataullakhanov FI McIntosh JR Nicastro D Cryo-fluorescence microscopy facilitates correlations between light and cryo-electron microscopy and reduces the rate of photobleaching J Microsc 2007 227 98 109 17845705
  22. [22]van Driel LF Valentijn JA Valentijn KM Koning RI Koster AJ Tools for correlative cryo-fluorescence microscopy and cryo-electron tomography applied to whole mitochondria in human endothelial cells Eur J Cell Biol 2009 88 669 684 19726102
  23. [23]Agronskaia AV Valentijn JA van Driel LF Schneijdenberg CTWM Humbel BM van Bergen en Henegouwen PMP Integrated fluorescence and transmission electron microscopy J Struct Biol 2008 164 183 189 18664385
  24. [24]Faas FGA Bárcena M Agronskaia AV Gerritsen HC Moscicka KB Diebolder CA Localization of fluorescently labeled structures in frozen-hydrated samples using integrated light electron microscopy J Struct Biol 2013 181 283 290 23261400
  25. [25]Le Gros MA McDermott G Uchida M Knoechel CG Larabell CA High-aperture cryogenic light microscopy J Microsc 2009 235 1 8 19566622 PMC3350647
  26. [26]Mastronarde DN Automated electron microscope tomography using robust prediction of specimen movements J Struct Biol 2005 152 36 51 16182563
  27. [28]O’Neil A Reichhardt C Johnson B Prevelige PE Douglas T Genetically programmed in vivo packaging of protein cargo and its controlled release from bacteriophage P22 Angew Chem Int Ed Engl 2011 50 7425 7428 21714051
  28. [29]O’Neil A Prevelige PE Basu G Douglas T Coconfinement of fluorescent proteins: spatially enforced communication of GFP and mCherry encapsulated within the P22 capsid Biomacromolecules 2012 13 3902 3907 23121071
  29. [30]Kukulski W Schorb M Kaksonen M Briggs JAG Plasma membrane reshaping during endocytosis is revealed by time-resolved electron tomography Cell 2012 150 508 520 22863005

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Authors
Schorb, M., & Briggs, J. A. G.
Affiliations
European Molecular Biology Laboratory (DE)
Corresponding author
Briggs, John A.G.
ORCID
Venue
Ultramicroscopy
Publisher
Elsevier BV
Published
2014-08
Volume
143
Pages
24-32
ISSN
0304-3991 (ISSN-L)
DOI
10.1016/j.ultramic.2013.10.015
PMC
PMC5472196
OpenAlex
W2053297551
Cited by
140
References
0
Citations by year
2026: 5 · 2025: 11 · 2024: 11 · 2023: 10 · 2022: 6 · 2021: 12 · 2020: 6 · 2019: 19 · 2018: 14 · 2017: 15 · 2016: 11 · 2015: 8 · 2014: 12
FWCI
5.17
Type
journal article
Language
en
Open access
Licence
Locations
  • Ultramicroscopy (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Advanced Electron Microscopy Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
  • Physical Sciences › Engineering › Biomedical Engineering › Advanced Materials Characterization Techniques
Keywords
high numerical aperture objective, cryo-EM sample preparation
MeSH terms
Bacteriophages, Fluorescent Dyes, Microscopy, Electron, Microscopy, Fluorescence, Sensitivity and Specificity, Cryoelectron Microscopy

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