CryoCLEM sample transfer solutions to enable sustainable use of cryo-microscopes

• Applicant: Dr. Roland Thünauer
• Subject Area: Structural Biology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 566291588

Correlative light and electron cryo-microscopy (cryoCLEM) offers the unique opportunity to unravel biological mechanisms by resolving protein structures directly in cells through combining the advantages of light microscopy with those of electron cryo-microscopy (cryoEM). Recent years have seen significant improvements in the available cryoEM and light cryo-microscopy technologies, resulting in the rapid introduction of new instruments by manufacturers. However, this has also led to the introduction of many different sample holder standards that are incompatible with equipment from competitors and/or older device generations. The integration of cryo-microscopes, which are typically very expensive and have a long lifetime, but rely on different sample holders, into a cryoCLEM workflow is possible but very difficult in practice due to the lack of solutions for safe, clean, and fast transfer of cryoCLEM samples between arbitrary sample holders. In addition, existing sample holder standards may limit the choice of new instruments that need to be integrated into a cryoCLEM workflow. Retrofitting the sample handling mechanics within existing cryo-microscopes to use new sample holders is not an option, as it is excessively laborious and usually cannot be done without intensive support from the microscope manufacturer. Therefore, we propose to develop a modular, open-platform cryo-sample transfer station that allows samples to be moved between different sample holders along the cryoCLEM workflow without the need to modify the microscopes themselves. The modular design of the transfer station will allow it to be customized and equipped with modules containing the docking mechanisms that can accommodate shuttles for the specific sample holder types that are required by a laboratory according to its cryoCLEM instrumentation. Furthermore, docking modules for sample transfer from other cryo-imaging modalities, such as X-ray cryo-microscopy, can also be integrated. The transfer station will be designed to minimize contamination during sample transfer and to allow safely transferring samples containing infectious pathogens. All design drawings and manufacturing instructions for the transfer station and docking modules will be made publicly available on a website. In addition, we will provide advice and support to interested laboratories and facilities so that they can customize and manufacture their own version of the transfer station. Taken together, the modular cryo-transfer station will enable cryoCLEM workflows that incorporate cryo-microscopes from different generations and manufacturers that rely on different sample holder types. This allows existing instruments to remain in operation longer and be used more efficiently in cryoCLEM workflows. In addition, new instruments with a different sample holder standard can be better integrated into existing cryoCLEM workflows.

• DFG Programme: Further Instrumentation Related Funding