Cryo Electron Microscopy – CryoEM

State-of-art cryo transmission electron microscope FEI Titan Krios 3Gi enables researchers to look at the macromolecules almost in their natural environment. Frozen hydrated specimens of e.g. proteins make high resolution structural analysis possible. Acquiring SPA datasets gives opportunity to get molecule structure with resolution of about 2Å, which means no more ‘blobology’ and very good insight in macromolecules internals. The use of direct electron detectors push scientific work output even further. Very fast data acquisition, with framerate up to 75 fps, drift correction and image averaging is now possible during the measurement itself. Such procedure drastically reduces disk size of datasets and allows to obtains more images at the same period of time comparing to work without direct detectors. Well known disadvantage of vitrificated samples imaging i.e. low contrast, can also be overcame with the use of direct electron detectors and when combined with usage of phase plate, macromolecular structures can be reconstructed reproducibly with very high, unreachable until now, resolutions and quality.

Staying in the subject of time, work efficiency and reproducibility requires a few word about automation to be said. Krios system is equipped with autoloader able to hold and handle twelve samples. Additionally the use of dedicated software for data acquisition, either for SPA or angle tomography, allows full automation of collection process.

• 300kV
• Ultra-stable
• Semi-automated
• Grid AutoLoader
• Three-condenser lens system
• Gatan K3 BioQuantum
• BioQuantum Energy filter and embedding
• Thermo Falcon3EC
• Ceta 16M
• Phaseplate
Gatan K3 BioQuantum with energy filter and BioQuantum Embedding:
• field of view 5760 x 4092 px with max image size 11520 x 8184 px Super-resolution
• GPU-based motion correction
• >1500 full fps; >75 fps to storage
• DQE for 300kV (200kV): Nq peak >0.83(0.87); 0.5Nq >0.53(0.53)
Falcon 3EC
• 4096 × 4096 px with 14 μm pixel size
• 40 fps to storage
• drift correction
• DQE at 0.5Nq >0.7
Ceta 16M Camera 300 kV
• 4096 × 4096 px CMOS sensor with 14 μm pixel size
• up to 25fps (512x512px)
• DQE at 0.5Nq >0.09

Support laboratory for sample preparation
1. Vitrification device
• The workstation is necessary to prepare frozen samples (below -170 degrees Celsius) of aqueous solution, without the formation of ice crystals within or on the surfaces of the respective samples.
• completely controlled digitally through a graphical user interface.
• the process carried out in the environmental chamber with the control of such parameters as: temperature: 5 to 60 degrees Celsius, humidity: up to 100%
• the possibility of freezing the preparation in liquid ethane or propane
• the possibility of automatic, simultaneous and bilateral blotting of grids (removal of excess material from dedicated TEM grid) and the regulation of its duration
• the device enable the user to repeat the process of simultaneous, double-sided drying of a single TEM mesh with a single blotting paper at least 16 times within one cycle.

2. Glow Discharger
• high-quality coater designed to produce homogeneous coatings of conductive metal or carbon as required for electron microscopy,
• instrument equipped with software enabling fully automated protocols
• combined configuration with interchangeable heads for sputter coating
• carbon thread evaporation coating
• all planetary rotation for even distribution of coating material
• glow discharge protocols to make TEM grids more hydrophilic

3. Isolation dewars – each for at least 400 grid boxes (4 grids each) plus holder system + high pressure thermal isolation dewars

IT infrastructure for data storage
• 72TB on storage with Falcon 3EC included
• additional storage server for 200 TB (enabling to store the data for 3 weeks after the

Sebastian Glatt

Detailed information can be found on the beamline’s main page.