Automated vitrification of cryo-EM samples with controllable sample thickness using suction and real-time optical...

Area of Science:

• Structural Biology
• Biophysics
• Microscopy Techniques

Background:

• Cryo-electron microscopy (cryo-EM) has seen rapid advancements in data collection and image processing.
• Cryo specimen preparation remains a bottleneck, necessitating faster and more reproducible methods.

Purpose of the Study:

• To present a novel, automated vitrification device for cryo-EM specimen preparation.
• To enable controlled and reproducible thin film preparation with integrated quality assessment.

Main Methods:

• Development of a vitrification device with automated sample handling and dew-point control.
• Removal of excess liquid via suction tubes, enabling light microscopy inspection.
• Vitrification of various biological samples including protein suspensions, lipid vesicles, and cells.

Main Results:

• The device allows for limited user interaction and reproducible thin film preparation.
• Integrated light microscopy enables quality assessment before electron microscopy data acquisition.
• Successful vitrification demonstrated for diverse biological samples.

Conclusions:

• The developed vitrification device significantly improves cryo-EM specimen preparation efficiency and reproducibility.
• Pre-imaging quality control enhances the reliability of cryo-EM data acquisition.
• The device is versatile, applicable to single particle analysis, cryo-electron tomography, and cryo-correlated light and electron microscopy.