Imaging intracellular components in situ using super-resolution cryo-correlative light and electron microscopy

Area of Science:

• Structural Biology
• Cell Biology
• Microscopy

Background:

• Super-resolution cryo-correlative light and electron microscopy (SRcryoCLEM) integrates cryo-single-molecule localization microscopy (cryoSMLM) and cryo-electron tomography (cryoET).
• This technique allows for precise localization and high-resolution imaging of biomolecules in their native cellular context.
• Challenges in SRcryoCLEM include specialized equipment and expertise requirements.

Purpose of the Study:

• To present a streamlined workflow for SRcryoCLEM.
• To demonstrate the integration of cryoSMLM into established cryoET protocols.
• To facilitate the adoption of SRcryoCLEM in structural biology research.

Main Methods:

• Implementation of a cryoSMLM workflow for precise biomolecule localization.
• Targeted cryo-electron tomography (cryoET) acquisition guided by cryoSMLM data.
• Correlation of cryoSMLM and cryoET datasets utilizing the scNodes software package.

Main Results:

• Successful execution of all SRcryoCLEM experiment stages using Vimentin as a target.
• Demonstration of cryoSMLM integration into a standard cryoET workflow.
• Validation of scNodes software for correlating cryoSMLM and cryoET data.

Conclusions:

• SRcryoCLEM is an accessible technique for in situ imaging of specific intracellular components.
• The presented workflow simplifies the process, making SRcryoCLEM more attainable.
• This method enhances the ability to study biomolecular structures within their cellular environment.