Name: Scanning transmissie-elektronenmicroscopietomografie in de virologie: 3D-beeldvorming van onder hoge druk ingevroren, vriesgesubstitueerde monsters (Video) | JoVE Journal
Uploaded: 2025-08-06T12:30:00.000+00:00
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Description: Leer STEM-tomografie voor virologisch onderzoek met hoge-druk invriezen en vriesubstitutie. Deze methode maakt 3D-beeldvorming van virusgeïnfecteerde cellen mogelijk voor inzicht in structurele veranderingen.

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</ol>

<table><tbody><tr><td>1-Hexadecene</td><td>Merck KGaA</td><td>8220640500</td><td></td></tr><tr><td>200 koperrooster met parallelle staven</td><td>Plano GmbH</td><td>G2014C</td><td></td></tr><tr><td>37 &deg;C droogkamer</td><td>Binder GmbH</td><td>9010-0323</td><td></td></tr><tr><td>Aceton</td><td>VWR international GmbH</td><td>20066330</td><td></td></tr><tr><td>DDSA</td><td>Electron Microscopy Sciences</td><td>13710</td><td></td></tr><tr><td>Diamantmes</td><td>Diatome Ltd</td><td>https://www.diatomeknives.com/ultra-35</td><td></td></tr><tr><td>DMP-30</td><td>Electron Microscopy Sciences</td><td>13600</td><td></td></tr><tr><td>Embed-812 Resin</td><td>Electron Microscopy Sciences</td><td>14900</td><td></td></tr><tr><td>EM tools</td><td>TVIPS GmbH</td><td>TVIPS Software: EM-Tools; https://www.tvips.com/imaging-software/em-tools/</td><td></td></tr><tr><td>Ethanol 100%</td><td>Merck KGaA</td><td>1.00983.2511</td><td></td></tr><tr><td>Filterpapier</td><td>VWR international GmbH</td><td>516-0812</td><td></td></tr><tr><td>Flessen met klikdekselglas 11 mL</td><td>VWR international GmbH</td><td>548-0625</td><td></td></tr><tr><td>Formvar</td><td>Electron Microscopy Sciences</td><td>15830-25</td><td></td></tr><tr><td>Freeze substitution unit Leica EM AFS2</td><td>Leica Microsystems GmbH</td><td>https://www.leica-microsystems.com/products/sample-preparation-for-electron-microscopy/p/leica-em-afs2/</td><td></td></tr><tr><td>Glod Sol 25 nm</td><td>AURION Immuno Gold Reagents &amp; Accessoires</td><td>425.011</td><td></td></tr><tr><td>Goudpinehole &oslash; 2000 &mu;m</td><td>Science Services GmbH</td><td>GA2000-Au</td><td></td></tr><tr><td>Gratiekels letters-netje koper</td><td>Plano GmbH</td><td>NH6C</td><td></td></tr><tr><td>Verwarmingsplaat</td><td>MEDAX GmbH &amp; Co. KG</td><td>12501</td><td></td></tr><tr><td>Hogedrukapparaat BAF 300</td><td>BAL-TEC AG</td><td></td><td></td></tr><tr><td>Hogedrukvriezer Wohlwend HPF Compact 01</td><td>Engineering Office M. Wohlwend GmbH</td><td>https://www.wohlwend-hpf.ch/index.html</td><td></td></tr><tr><td>Vloeibare stikstoftank</td><td>H.Erben GmbH</td><td>94500</td><td></td></tr><tr><td>Stikstof (vloeibaar)</td><td>VWR international GmbH</td><td>-</td><td></td></tr><tr><td>NMA</td><td>Electron Microscopy Sciences</td><td>19000</td><td></td></tr><tr><td>Osmiumtetraoxide</td><td>ChemPur Feinchemikalien und Forschungsbedarf GmbH</td><td>006051</td><td></td></tr><tr><td>Oven 120 &deg;C</td><td>Binder GmbH</td><td>9010-0194</td><td></td></tr><tr><td>PELCO easi-Glow</td><td>Ted Pella, Inc.</td><td>91000</td><td></td></tr><tr><td>Poly-L-Lysine 0,1% w/v aq. oplossing</td><td>Ted Pella, Inc.</td><td>18026</td><td></td></tr><tr><td>Safe-Lock Tubes 0,5 mL</td><td>Eppendorf SE</td><td>0030 121.023</td><td></td></tr><tr><td>Staalbewerkingscontainers (Beem capsules)</td><td>Plano GmbH</td><td>G360-1</td><td></td></tr><tr><td>Saffier schijf dia. 3 x 0,16 mm</td><td>Engineering Office M. Wohlwend GmbH</td><td>500</td><td></td></tr><tr><td>Ultramicrotome Leica EM UC7</td><td>Leica Microsystems GmbH</td><td>https://www.leica-microsystems.com/de/produkte/em-probenvorbereitung/p/leica-em-uc7/</td><td></td></tr><tr><td>Uranyl acetate</td><td>Riedel de Ha&euml;n AG</td><td>31697</td><td></td></tr><tr><td>UV lichtkamer</td><td>Dinies</td><td>ELG100S</td><td></td></tr></tbody></table>

scanning transmissie-elektronenmicroscopietomografie in de virologie: 3d-beeldvorming van onder hoge druk ingevroren, vriesgesubstitueerde monsters

Scanning Transmission Electron Microscopy Tomography in Virology: 3D Imaging of High-pressure Frozen, Freeze-substituted Samples

Scanning transmissie-elektronenmicroscopietomografie in de virologie: 3D-beeldvorming van onder hoge druk ingevroren, vriesgesubstitueerde monsters

Electron microscopy (EM) and especially three-dimensional (3D) EM techniques have become established methods in structural virology. Investigation of ...

scanning-transmission-electron-microscopy-tomography-virology-3d

Research

JoVE Journal

Medicine

Geneeskunde

Het protocol beschrijft een high-throughput benadering voor het bepalen van de structuren van membraaneiwitten met behulp van cryo-electron tomografie-en 3D-beeldverwerking. Het omvat de details van prepareren, data verzamelen, data verwerken en interpreteren, en afgesloten met het vervaardigen van een vertegenwoordiger doelstelling voor de aanpak, de HIV-1 Envelop glycoproteïne. Deze computationele procedures zijn ontworpen op een manier die onderzoekers en studenten in staat stelt om op...

Bepaling van de moleculaire structuren van HIV envelop glycoproteïnen met behulp van Cryo-electron tomography en geautomatiseerde Sub-tomogram Middeling

We beschrijven een correlatieve microscopie methode die high-speed 3D levende cellen fluorescerend licht microscopie en hoge-resolutie cryo-electron tomografie combineert. We demonstreren het vermogen van de correlatieve wijze dynamische beeldvorming, kleine HIV-1 deeltjes interactie met gastheer HeLa cellen.

Correlatieve microscopie voor 3D Structural Analysis van Dynamic Interactions

We presenteren een protocol over hoe om te verzamelen en te verwerken elektronen cryo-tomogrammen van hele mitochondriën. De techniek biedt gedetailleerd inzicht in de structuur, functie en organisatie van grote membraan eiwitcomplexen in de moedertaal van biologische membranen.

Visualisatie van ATP synthase Dimeren in Mitochondriën door Electron Cryo-tomografie

De bevestiging van virionen aan een oppervlak is een vereiste voor enkelvoudige virion imaging door Super-resolution fluorescentie imaging of atomic force microscopie (AFM). Hier demonstreren we een monstervoorbereidingsmethode voor gecontroleerde hechting van virionen op glasoppervlakken die geschikt zijn voor gebruik in AFM- en superresolutiefluorescentiebeeldvorming.

Monstervoorbereiding voor single virion atomic force microscopie en superresolutie fluorescentie beeldvorming

Dit rapport beschrijft een monstervoorbereidingsprotocol en specifieke beeldvormingscondities voor het uitvoeren van schuifende transmissie-elektronentomografie van dikke biologische monsters.

Voorbereiding en observatie van Thick biologische monsters door Scanning Transmission Electron Tomografie

Dit protocol wordt beschreven hoe de voorbijgaande verdichting van het DNA in cyanobacteriën visualiseren. Synchrone teelt, toezicht door fluorescentie microscopie, snelle invriezen, en hoge spanning cryo-Elektronentomografie worden gebruikt. Een protocol voor deze methodologieën wordt gepresenteerd, en toekomstige toepassingen en ontwikkelingen worden besproken.

Visualisatie van DNA verdichting in cyanobacteriën door hoogspannings-Cryo-Elektronentomografie

Hier wordt een protocol gepresenteerd dat de monstervoorbereiding en gegevensverzamelingsstappen beschrijft die nodig zijn in cryo zachte röntgentomografie (SXT) om de ultrastructuur van hele cryo-geconserveerde cellen in beeld te brengen met een resolutie van 25 nm halve toonhoogte.

Een 3D cartografische beschrijving van de cel door Cryo Soft X-ray Tomography

We presenteren een protocol voor lamellenpreparaat van diepgevroren biologische monsters door cryo-gerichte ionenbundelmicromachining voor structurele studies met hoge resolutie van macromoleculen in situ met cryo-elektronentomografie. Het gepresenteerde protocol biedt richtlijnen voor de bereiding van hoogwaardige lamellen met een hoge reproduceerbaarheid voor structurele karakterisering van macromoleculen in de Saccharomyces cerevisiae.

Bereiding en Cryo-FIB microbewerking van Saccharomyces cerevisiae voor Cryo-Elektronen Tomografie

Hier worden protocollen beschreven om virusassemblages voor te bereiden die geschikt zijn voor vloeistof-EM- en cryo-EM-analyse op nanoschaal met behulp van transmissie-elektronenmicroscopie.

Geavanceerde beeldvorming met hoge resolutie van virusassemblages in vloeistof en ijs

Cryo-STEM-tomografie biedt een middel om organellen van intacte cellen te visualiseren zonder inbedding, sectie of andere invasieve preparaten. De verkregen 3D-resolutie ligt momenteel in het bereik van enkele nanometers, met een gezichtsveld van enkele micrometers en een toegankelijke dikte in de orde van grootte van 1 μm.

Visualisatie van organellen in situ door cryo-STEM tomografie

Leer STEM-tomografie voor virologisch onderzoek met hoge-druk invriezen en vriesubstitutie. Deze methode maakt 3D-beeldvorming van virusgeïnfecteerde cellen mogelijk voor inzicht in structurele veranderingen.

Scanning transmissie-elektronenmicroscopietomografie in de virologie: 3D-beeldvorming van onder hoge druk ingevroren, vriesgesubstitueerde monsters (Video) | JoVE Journal

Scanning transmissie-elektronenmicroscopietomografie in de virologie: 3D-beeldvorming van onder hoge druk ingevroren, vriesgesubstitueerde monsters

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