Linkam presents cryo-plunging techniques at Microscopy Conference 2023

Linkam presents cryo-plunging techniques for automated vitrification of cryo-electron microscopy samples at Microscopy Conference 2023, 26^th February – 2^nd March

02 March 2023: Linkam Scientific will present a new cryo-electron microscopy (cryo-EM) research paper at the Microscopy Conference 2023 (MC2023), Darmstadt, Germany: ‘New principle and robot for vitrification of cryo-EM samples with controllable sample thickness and real-time optical inspection’. Lead author of the paper, Linkam’s senior research scientist Dr. Michael Schwertner, will present new findings in cryo-EM at 12:15pm (CET) on 1^st March within the MC2023 Instrumentation and Methods session, in the Development of Cryo-EM Instrumentation and Techniques track.

The paper explores how, despite recent progress in cryo-EM instruments to support automated sample loading and imaging, the routine preparation of cryo samples still presents a challenge. Current cryo-EM methods are held back by the speed and quality of sample preparation because of the difficulty of creating a uniform film, and a more robust and reproducible method has long been sought by the research community. The paper presents that utilising suction rather than blotting paper to adjust the film thickness before plunging can improve sample preparation, by removing the potential for the blotting paper to create a challenging environment inside the high humidity chamber. It also gives access for real-time imaging that is used to assess sample conditions, and to determine the best moment to trigger plunging.

Linkam’s new generation cryo plunger, the CryoGenium, is a highly automated device that prepares samples for cryo imaging by plunge-freezing via liquid ethane,^[i] a technique used to flash freeze biological specimens. Cryo fixation, which involves plunging a sample grid into vitreous water, delivers structural preservation of biological samples at atomic resolution, including protein suspensions, viruses, bacteria and eukaryotic cells. Plunge freezing at >100.000^oC/s prevents ice crystals forming in the sample in the vitrification process.^[ii]

The plunge-freezing device workflow automates steps in the sample preparation process, including grid handling, glow discharging, controlling cryogenic liquids, and sample application. Because it uses sample removal by suction rather than filter paper, and allows visual inspection of the grid during thin film formation, it enables precise timing of vitrification by the user.

Dr. Schwertner shares an insight into his presentation: “By comparing traditional cryo sample preparation methods with our new approach of sample application via dipping, followed by film thickness adjustment via suction, researchers are able to optimise cryo-EM sample preparation. We envisage that future decisions on the precise moment to plunge the sample will be automated via an artificial intelligence (AI) vision system.”[iii]  

The Linkam CryoGenium can be used in the analysis of protein suspensions, lipid vesicles, bacteria and human cells. Prototypes of the CryoGenium are currently undergoing field trials prior to launch. To talk to Linkam about its new portfolio of cryo-EM solutions and sample characterisation methods and to meet us at the show, contact our team of experts at info@linkam.co.uk.

About Linkam Scientific Instruments

Linkam develops and manufactures a broad range of temperature and environmental control stages for both OEMs and end users. From high to cryo temperatures as well as humidity, electrical connections, gas purging, vacuum and pressure, for enhanced sample analysis. Linkam stages are used with light microscopes and a wide range of analytical techniques including Raman, FTIR, SAXS-WAXS and many more to visualise and characterise the properties of materials. Linkam stages are found in thousands of laboratories worldwide with the most successful microscope heating stage, the THMS600, selling over 6,000 units alone. Linkam is the market leader in temperature-controlled microscopy.  

REFERENCES

[i] Tivol, W. F., Briegel, A. & Jensen, G. J. An improved cryogen for plunge freezing. Microsc. Microanal. 14, 375–379 (2008)

[ii] Koning, R.I., Vader, H., van Nugteren, M. et al. Automated vitrification of cryo-EM samples with controllable sample thickness using suction and real-time optical inspection. Nat Commun 13, 2985 (2022). https://doi.org/10.1038/s41467-022-30562-7

[iii] Koning, R.I., Vader, H., van Nugteren, M. et al. Automated vitrification of cryo-EM samples with controllable sample thickness using suction and real-time optical inspection. Nat Commun 13, 2985 (2022). https://doi.org/10.1038/s41467-022-30562-7