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Thermo Scientific Krios


CEMAS offers access to a comprehensive suite of instruments needed for cryo-electron microscopy (cryoEM) research that encompasses the entire workflow from sample preparation to data analysis. This powerful technique allows for the observation of biological specimens in their native environment by embedding the samples in vitreous ice. Recent advances in camera technology and data processing have made it possible to determine molecular structures at near-atomic resolution. CryoEM has transformed structural biology, and its use is expanding into other fields such as small molecule crystallography, infectious disease, and cell biology.

In the spring of 2019, CEMAS installed the Thermo Scientific Krios G3i cryoEM which has a 300 kV x-FEG source and can be configured for single particle analysis (SPA), tomography, and other applications.

Krios capabilities include:

 

Krios microscopeHigh resolution performance

The Krios cryoEM has a proven track record of producing high resolution molecular structures including the majority of published structures below 4 Å. These high-resolution structures were achieved in part because of the system’s stability (reduced thermal drift) during data acquisition sessions. The microscope is configured with a three-condenser lens system for continuous, parallel sample illumination.

Gatan BioQuantum energy filter and K3 camera

The Krios is equipped with a post-column Gatan BioQuantum energy filter that removes inelastically scattered electrons before reaching the detector. This filter enhances contrast in the image, which can improve particle alignment and overall map resolution.

The energy filter is paired with the newest direct electron detector from Gatan – the K3 camera. Both the energy filter and camera are suited for low-dose imaging applications with the highest DQEs available. The K3 sensor has a field of view of 24 megapixels (5,760 x 4,092), which is 1.6 times the size of the K2 camera. Additionally, the K3 can capture 1,500 full frames per second (3.75 times the speed of the K2 camera).

Additional configurations

Included on the Krios is a phase plate solution that enhances the contrast of small particles (below 200 kDa) without having to work at large defocus values, thereby compromising low resolution information. This will be particularly beneficial for those interested in cryo-electron tomography as it allows each tilt image to be collected at a lower electron dose and reduces sample damage.

This microscope is also equipped with a Cs image corrector that not only minimizes the spherical aberration but other common aberrations.

Workflow connectivity

Like the Glacios, the Krios is equipped with an Autoloader system, a robotic sample handler, which can hold up to 12 grids per batch. It is expected that users will pre-screen their samples on the Glacios before placing them in the Krios. Because of the compatibility of the Autoloaders, samples can be easily transferred from the Glacios to the Krios without contamination.

Enhanced ease-of-use through automation 

EPU software is the native software package for single particle data acquisition (separate tomography software is available). EPU is capable of batch-screening samples and categorizes grids by ice thickness to further expedite data collection. The program automates data collection from different grid squares and optimizes conditions before collecting images.