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Cryo-electron tomography (cryo-ET) provides unprecedented insights into the inner workings of cells, but clear, reliable results depend on high-quality cryo-lamella preparation. The Thermo Scientific Arctis Cryo-Plasma Focused Ion Beam (Cryo-PFIB) is specifically designed for automated, high-throughput production of cryo-lamellae from vitrified cells. Its Autoloader system provides a unique, direct connection between cryo-FIB-SEM sample preparation and cryo-transmission electron microscopy (cryo-TEM) within the tomography workflow.
Watch on demand to learn more about the Arctis Cryo-Plasma-FIB, including:
An "on-board" integrated wide-field fluorescence microscope (iFLM) allows the same sample area to be observed with light, ion, or electron beams. Specially designed TomoGrids ensure correct cryo-lamellae alignment to the tomographic tilt axis, from initial milling through high-resolution TEM imaging.
The iFLM Correlative System enables fluorescence imaging at the electron/ion beam coincidence point. Switching between iFLM targeting and ion milling can be performed without moving the stage. The 180° tilt capability of the CompuStage allows for imaging of the top and bottom surfaces of the sample, which can be helpful for thick samples.
TomoGrids are specifically made for the cryo-tomography workflow and have two flat sides facing each other. These sides prevent misalignment during loading into the cryo-TEM and always ensure proper orientation of the lamella axis with respect to the TEM tilt axis. With TomoGrids, the entire available lamella area can be utilized for data acquisition.
The Arctis Cryo-PFIB maintains an ultra-clean working environment for several days, ensuring consistent, high-quality lamella preparation. The plasma ion-beam source can switch between xenon, oxygen, and argon ions, facilitating the production of very thin lamellae with optimized surface quality.
PFIB technology enables applications that are not covered by liquid metal ion source (LMIS) FIB systems. For example, it is possible to use the different milling properties of the three available ion beams to produce a high-quality sample while avoiding gallium implantation effects. System enclosure is designed with biosafety in mind and an a heat decontamination solution is optionally available for higher biosafety level labs (e.g. BSL 3).
The compact sample chamber of the Arctis Cryo-PFIB is specifically designed for cryo-operation. Its reduced volume allows for an exceptionally clean operating environment that minimizes water condensation. This designed-in cleanliness is further enhanced by sample cooling via braids and sample shielding with a dedicated cryo-box, ensuring a working environment that routinely allows for multi-day batch sample preparation.
The Autoloader enables robotic sample loading and unloading for up to 12 grids (TomoGrids or AutoGrids), facilitating transfer to the cryo-TEM while minimizing sample damage and contamination risks. In the new web-based user interface loaded grids are first mapped and inspected. Subsequently, lamella positions are chosen, and milling parameters are defined. Milling work is carried out autonomously. Depending on the specimen, the plasma source enables high milling rates for fast, large-volume material removal.
The Autoloader cassette provides a protected environment for the delicate cryo-lamellae samples. Risky manual sample manipulation steps that could damage or contaminate the samples are largely avoided. The cassette is loaded into a capsule docked to the Autoloader, which can be interchanged between the Arctis Cryo-PFIB and Krios or Glacios Cryo-TEMs.
"One of the things that I find exciting about Arctis [Cryo-PFIB] is the degree to which it is an automated workflow. And so, this ability to have it running essentially unsupervised and produce high quality, large numbers of lamella is transformational."
Jim Naismith, Director of The Rosalind Franklin Institute and key collaborator on the development of the Thermo Scientific Arctis Cryo-PFIB.
Thermo Fisher Scientific and The Rosalind Franklin Institute have been working together since 2019 to determine what’s next for cryo-EM. Through this partnership, we developed and built the Thermo Scientific Arctis Cryo-PFIB.
Cryo-electron tomography delivers both structural information about individual proteins as well as their spatial arrangements within the cell. This makes it a truly unique technique with enormous potential for cell biology; cryo-electron tomography is bridging the gap between light microscopy and atomic-resolution techniques like single particle analysis.
Ion optics | |
|---|---|
| Ion gun | High-performance PFIB column, with inductively coupled plasma (ICP) source supporting fast ion switching |
| Species | Xenon, argon, oxygen |
Nitrogen cooling and runtime | |
|---|---|
| Nitrogen refilling | Automatic liquid nitrogen filling system |
| Cryo-runtime | Up to multiple days |
Electron optics | |
|---|---|
| Electron gun | High-stability Schottky field emission gun |
Conductive coating | |
|---|---|
| Sputtering | Sputter target for conductive sputter coating |
Fluorescence microscope | |
|---|---|
| Coincidence point | Triple beam coincidence at sample position for photons, electrons, and ions |
Protective coating | |
|---|---|
| Gas injection system | Retractable platinum GIS for chemical vapor deposition |
Stage and sample loader | |
|---|---|
| Stage | Cryo-CompuStage |
| Sample loader | Autoloader: automated loading of cassettes (up to 12 AutoGrids/TomoGrids) under cryo-conditions |
Environmental protection | |
|---|---|
| Enclosure | System fully enclosed designed with biosafety in mind |
Cryo-EM can determine the structural features of protein aggregates implicated in neurodegenerative diseases, allowing scientists to address how they form, interact with the cellular environment, and alter brain function.
Cryo-Tomography
Cryo-electron tomography (cryo-ET) delivers both structural information about individual proteins as well as their spatial arrangements within the cell. This makes it a truly unique technique and also explains why the method has such an enormous potential for cell biology. Cryo-ET can bridge the gap between light microscopy and near-atomic-resolution techniques like single-particle analysis.
MicroED
MicroED is an exciting new technique with applications in the structural determination of small molecules and protein. With this method, atomic details can be extracted from individual nanocrystals (<200 nm in size), even in a heterogeneous mixture.
Cryo-Tomography
Cryo-electron tomography (cryo-ET) delivers both structural information about individual proteins as well as their spatial arrangements within the cell. This makes it a truly unique technique and also explains why the method has such an enormous potential for cell biology. Cryo-ET can bridge the gap between light microscopy and near-atomic-resolution techniques like single-particle analysis.
MicroED
MicroED is an exciting new technique with applications in the structural determination of small molecules and protein. With this method, atomic details can be extracted from individual nanocrystals (<200 nm in size), even in a heterogeneous mixture.
Cryo-EM enables the 3D structural visualization of virus particles, and the antigen-antibody interface, at near-atomic resolutions. A virus’s inherent structural symmetry makes it the ideal target for cryo-EM analysis.
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