Cryo-electron Microscopy and Tomography

Modern electron microscopy in structural biology on the cellular and molecular level is performed by cryo-electron microscopy and cryo-electron tomography. Cryo-electron tomography (cryo-ET) is the only method to address pleiomorphic structures like cells and organelles in a close to native state, while cryo-electron microscopy (cryo-EM) is applied to study single particles, primarily larger macromolecular complexes, which have been isolated and purified by biochemical methods. Both methodologies provide information on the cellular and molecular level and are therefore ideal for in-depth structural-functional analysis in combination with state of the art biochemical characterisation.

Services provided: Cryo-electron microscopy and tomography core facility CEITEC MU provides access and support with acquisition of cryo-electron microscopy images for both single particle as well as for cryo-electron tomography applications. Furthermore, the facility provides support with sample preparation, namely plunge freezing of in vitro purified samples, as well as cryo-FIB lamella micromachining of cells. Additional support is given in experiment design, setting up data collection strategy and data analysis (i.e. image processing). Interested non-specialists can receive training in operation of the electron microscopes or develop a collaboration with Cryo-electron microscopy and tomography core facility CEITEC MU, particularly in these areas: high-resolution structure determination of macromolecular complexes and assemblies, structural studies of intracellular compartments and host-pathogen interactions, time-resolved electron microscopy of transient macromolecular complexes.

Key Equipment

  • High-end electron microscope for high-resolution cryoEM and cryo-tomography FEI Titan Krios (80 - 300 kV) equipped with an energy filter and a direct detector camera
  • Conventional cryoEM microscope FEI F20 (200 kV) equipped with a CCD camera and a dual beam FIB/SEM instrument (Versa3D) for micromachining of thin lamellas of vitrified cells usable for electron tomography
  • Equipment for sample preparation
  • Vitrification robot Vitrobot Mark IV
  • Thermo Scientific Talos Arctica – mid-range transmission electron microscope operating at 40 - 200 kV equipped with Schottky field emission gun (XFEG) providing highly coherent electron beam with ultimate stability. The microscope is equipped with Thermo Scientific Falcon3 EC direct electron detector and Ceta camera. Automated collection data for single particle analysis and cryo-electron tomography data can be performed with EPU or SerialEM software, respectively. The microscope is well suited for automated acquisition of high-quality cryoEM data, efficient screening of cryoEM samples, and 3D electron diffraction tomography

Expertise

  • Studies, at the highest currently possible level, of cellular structures and organelles, protein complexes and various phenomena at the macromolecular level by 3D imaging

Open-access

  • Advanced and routine application of cellular electron tomography
  • Application of correlative methods integrating light/fluorescence microscopy with high resolution cryo-electron microscopy techniques
  • Single particle analysis
  • Preliminary negative stain/RT- and advanced cryo-electron microscopy methods

Core Facility contact

Jiří Nováček - jiri.novacek@ceitec.muni.cz

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More information about Cryo-electron Microscopy and Tomography and related services you can find in document Equipement and services of CIISB (page 17).


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