Techniques available
Techniques that are currently available at the facility are:
- Standard 2D projection imaging of thin (cryo-) samples, providing structural information with high resolution.
- Single-particle analysis. Provides high-resolution structures of a single protein or protein complex. Single-particle analysis is the core strength of the facility. First, the quality of the sample is assessed with negative stain imaging and 2D class averages are obtained, after which the sample is vitrified and screened until the right conditions are reached to collect data. The data is then processed to produce a 3D structure of the particle. When necessary, samples can be taken to national facilities, such as the SCMI or eBIC.
- (Cryo-)electron tomography. Provides 3D information of a single specimen by physically tilting the specimen inside the EM. Can be used on cryo-samples as well as plastic sections of cellular material.
- Electron Diffraction. Provides structural information on crystalline material.
Equipment
- Thermo Fisher Scientific Tecnai F20 electron microscope (200 kV, field emission gun) equipped with an 3K CMOS camera (Gatan Rio) and a direct electron detector (Gatan K2 Summit).
- Single tilt liquid nitrogen cryo-transfer holders (Gatan 626, Gatan ELSA), with transfer-station, pumping station and temperature control unit.
- Thermo Fisher Scientific Vitrobot automated vitrification unit (Vitrobot mark IV).
- Various supporting equipment, such as a glow-discharge unit, negative stain facilities and a room-temperature holder.
- Extensive computer resources for data processing.
Training
The CryoEM facility provides extensive training for users, ranging from negative stain sample preparation, conventional room-temperature TEM, to plunge-freezing, cryo-TEM, and data-processing of single-particle datasets.