Cryo-Electron Microscopy

Advances in the detector technology and image processing have revolutionized electron microscopy. It is now a method of choice for structural characterization of protein complexes over 150 kDa. Both the resolution and the minimum size are likely to be improved further in the near future. In cell biology, tomography-based approaches can be applied to visualize subcellular structures with unprecedented resolution.

We have established capability to prepare and screen samples to a high standard. We also have computing infrastructure and expertise that can be used to support the processing of datasets obtained. We aim to build capability and we will endeavour to support all new projects. A combination of external courses and hands on training will be provided that ultimately enables researchers to operate independently.

Microscope:

Dundee electron microscopy facility hosts a JEOL JEM-2200FS electron microscope, equipped with a 200kV field emission gun (FEG) and an integrated in-column energy filter (omega filter). The microscope is currently equipped with a Gatan 4kx4k ultrascan CCD camera.

Capabilities:

JEOL JEM-2200FS is routinely used for single particle imaging. Images can be obtained with defined defocus using SerialEM acquisition software. High angle specimen tilt (+ 70 to -70) over a wide angular range can be obtained which enable tomographic reconstruction. Although the microscope is equipped for cryo applications, it can also be used as a standard TEM, for example during the early stages of sample optimization using negative stain EM.

Sample preparation:

We have a dedicated rapid plunge freezing device (vitrobot MarkIV) that allows preparation of particles embedded on vitrifed ice on a cryo grid suitable for single particle imaging. A glow discharger for cleaning and charging the grids is also available.

Access to National 300 keV Cryo-EM Facilities

In most cases, for collection of datasets at ultimate resolution, it is preferable to make use of 300 keV microscopes equipped with state-of-the art direct electron detectors. The EM facility in Dundee provides the opportunity to screen and optimise samples to ensure only highest quality samples are submitted to microscopes at National facilities.  These facilities include:

• UK national facility at eBIC (eBIC access for Scotland BAG currently led by Tom Owen-Hughes) 
• Scottish national facility, SCMI located in Glasgow (applications to be routed via the Dundee SCMI representative Tom Owen-Hughes). Costs for access to SMIC are £2177 per two-day session.

To access the national facilities please contact Professor Tom Owen-Hughes: t.a.owenhughes@dundee.ac.uk