Automated LN₂ refill device for uninterrupted cryoFIB-SEM operations

Area of Science:

• Structural biology
• Microscopy
• Biophysics

Background:

• In situ structural cell biology is advancing due to faster, more reliable workflows.
• Cryo-focused ion beam scanning electron microscopy (cryoFIB-SEM) is crucial for preparing biological samples for cryo-electron tomography (cryo-ET).
• Automated cryoFIB-SEM requires a continuous supply of liquid nitrogen (LN₂) for stable cryogenic conditions.

Purpose of the Study:

• To introduce a custom automated liquid nitrogen (LN₂) refill system for cryoFIB-SEM.
• To enable long-term, automated, and unsupervised lamellae preparation.
• To reduce manual labor and enhance operational efficiency in cryogenic microscopy.

Main Methods:

• Development of a custom automated LN₂ refill system.
• Integration with gas-cooled cryostages for cryoFIB-SEM instruments.
• Testing for compatibility with cryoSEM and cryoFIB-SEM devices requiring nitrogen gas-flow cooling.

Main Results:

• The automated LN₂ refill system ensures a constant supply of LN₂, maintaining cryogenic conditions.
• The system supports long-term cryoFIB-SEM operations, enabling unsupervised sample preparation.
• The solution effectively liberates users from repetitive manual tasks.

Conclusions:

• This automated LN₂ refill system significantly improves the efficiency and reliability of cryoFIB-SEM workflows.
• The system is beneficial for high-throughput structural cell biology and can be adapted for various cryogenic electron microscopy setups.
• It offers particular advantages in high-containment laboratories (BSL-3/BSL-4) by minimizing user presence.