Cryo-electron tomography on focused ion beam lamellae transforms structural cell biology

Area of Science:

• Structural Biology
• Cell Biology
• Microscopy

Background:

• Macromolecular structures are often studied in isolation, losing context of the cellular environment.
• In situ structural information is crucial for understanding native molecular functions and interactions.
• Cryo-electron microscopy (cryo-EM) provides high resolution but typically requires purified samples.

Purpose of the Study:

• To review recent advancements in cryo-electron tomography (cryo-ET) for in situ structural studies.
• To highlight the role of focused ion beam (FIB) fabrication in preparing cellular samples for cryo-ET.
• To discuss technological developments enabling higher resolution cryo-ET of cellular interiors.

Main Methods:

• Cryogenic focused ion beam (FIB) milling to create thin lamellae of cells and tissues.
• Cryo-electron tomography (cryo-ET) for high-resolution 3D reconstruction of cellular structures.
• Advancements in sample freezing, tomography, data processing, and correlative light and electron microscopy.

Main Results:

• Cryo-ET on FIB-prepared lamellae enables near-native imaging of macromolecular structures within cells.
• Technological progress has led to increasingly higher resolution cryo-ET data.
• In situ structural studies are revealing previously inaccessible molecular interactions.

Conclusions:

• Cryo-ET combined with FIB fabrication is a powerful tool for in situ structural biology.
• Continued technological development promises further breakthroughs in understanding cellular architecture and function.
• This approach is expanding the scope of structural studies to complex cellular environments.