Affiliated School or College: School of Medicine
Cryo-Electron Microscopy (Cryo-EM) is an approach that allows the observation of hydrated biological specimens in their native environment at cryogenic temperatures in transmission electron microscopy (TEM). Cryo-EM broadly encompasses three different approaches: electron crystallography, single-particle Cryo-EM, and Cryo-ET. Recent technological advancements in cryo-EM have ushered in a new era of structural biology enabling exclusive views of the complex biological molecules. Atomic details of biomolecules allows us to better understand physiological phenomena that govern life and aid in drug design and development.
Our facility focuses on cryoEM/ET for biological samples, along with conventional techniques such as ultramicrotomy and immunocytochemistry. The facility currently has four shared TEM microscopes and one cryo focussed electron beam- scanning electron microscope (cryo-FIB-SEM):
- Titan Krios G3i 300 kV X-FEG with BioQuantum K3, and Volta Phase Plate.
- Glacios 200 kV X-FEG with Seletrix Energy filter and Falcon IV DED camera.
- Aquilos II Cryo-FIB with CLEM system is the latest generation of our cryo-DualBeam system.
- Tecnai G² Spirit T12 (120kV) with a 4k x 4k Gatan US4000 CCD camera,
- Tecnai TF20 (200kV, FEG) with a 4k x 4k Tietz F416 CMOS and a DE-20 DED camera."
Organization Affiliations: Cleveland Center for Membrane and Structural Biology (CCMSB)
Customers: External and Internal
Hours
24/7
Location
Cleveland Center for Membrane and Structural Biology (CCMSB)
1819 E 101st Street
Cleveland, OH 44106
Rates
The facility is set up to study flash-frozen samples by Cryo-EM Single-Particle Analysis (SPA) or Cryo-electron Tomography (Cryo-ET). We welcome trained EM users, as well as researchers who wish to be trained. Several collaboration opportunities are available with CWRU School of Medicine faculty. The facility also offers complete conventional transmission electron microscopy (TEM) services, including: sample processing, ultramicrotomy, film developing, and photographic printing, as well as immunocytochemistry.
Services
- Single particle: sample evaluation, grid freezing, grid screening, data collection
- Single-particle cryo-EM can provide structural information for a large variety of biological molecules without the need to produce crystals. Proteins from 40 kDa to several MDa in size can be studied by this methodology. Very little sample is required for this technique. For cryo-EM the specimen, typically a purified protein sample, is embedded in vitreous ice on a cryo-EM grid, and is kept at cryogenic temperatures while images are recorded by the electron microscope. After completing the imaging, single-particle reconstruction methods are used to solve the structure of the protein. Current resolution for single particle cryo-EM is in the 1.5~3.5 Å range.
- Cryo-ET: sample evaluation, cryo-FIB, grid screening, data collection
- Cryo-ET can unveil the detailed 3D structures of cellular and subcellular macromolecular objects. High pressure-freezing techniques for cells and tissue can provide exceptional preservation of 100µm large samples in the native state. Single cells can be vitrified by plunging them into liquid ethane. This allows studying macromolecular complexes in their cellular environment, which provides a deep insight into cellular processes at the molecular level. 2D images (projections) of the sample collected at different angles allow the 3D structure to be reconstructed by the back-projection method. Resolution for cryo-electron tomography is in the 4-30 Å range.
- Conventional TEM service: negative staining,ultramicrotomy,immunocytochemistry.
- Consultation and Training: Consultation will cover the whole workflow and the training will cover any combination of the techniques listed above.
Pricing
Please check the “schedule equipment” page in iLab for instrument usage rate and the “request service” page for the consumable parts rate and staff assistance fee.