Direct deposition on pre-cooled grids: a new approach to sub-millisecond cryoEM grid preparation

Area of Science:

• Structural Biology
• Biophysics
• Microscopy

Background:

• Single-particle cryo-electron microscopy (cryo-EM) requires efficient sample preparation.
• Current methods face challenges with air-water interface interactions and speed limitations.
• Time-resolved studies are crucial for observing dynamic biological processes.

Purpose of the Study:

• To develop a significantly faster cryo-EM grid preparation technique.
• To overcome limitations associated with traditional plunging methods.
• To enable time-resolved studies on the millisecond timescale.

Main Methods:

• Developed a novel method involving direct spraying of samples onto pre-cooled grids within the cryogen.
• Eliminated the conventional plunging step in sample preparation.
• Achieved sample freezing in under 1 millisecond.

Main Results:

• Demonstrated a proof of principle for the direct spraying technique.
• Provided a detailed description of the methodology and its implementation.
• Successfully prepared grids with two mixed and frozen samples.

Conclusions:

• The new method drastically reduces sample preparation time for cryo-EM.
• This advancement opens new possibilities for time-resolved EM studies.
• The technique has the potential to improve sample quality and expand accessible research areas.