What Is Cryo-Electron Tomography?
Cryo-ET (also called TOMO or cryo-tomography) is a cryo-EM technique that produces 3D reconstructions of cellular environments by tilting the sample and collecting a series of images (a tilt series). Unlike SPA, which averages thousands of identical particles, tomography captures unique, one-of-a-kind structures in their native context.
How It Works
- Vitrification: Cells or tissue are rapidly frozen (high-pressure freezing or plunge freezing)
- Thinning: For cells >500nm thick, FIB-SEM is used to mill thin lamellae (~200nm)
- Tilt Series Collection: The sample is tilted from -60° to +60° in 2-3° increments
- Reconstruction: Alignment and back-projection produce a 3D tomogram
- Subtomogram Averaging: Identical particles within tomograms are extracted and averaged for higher resolution
- High-pressure freezing and FIB-SEM lamella preparation
- Tilt series collection on 300kV Titan Krios
- Tomogram reconstruction and subtomogram averaging
- HOPE-SIM cryo-fluorescence for targeted tomography
Cryo-ET vs SPA
| Feature | SPA | Cryo-ET (TOMO) |
|---|---|---|
| Sample | Purified protein | Intact cells/tissue |
| Context | In vitro | In situ (native) |
| Resolution | 1.4-4Å | 20-50Å (subtomogram: 5-15Å) |
| Throughput | High | Lower |
| Heterogeneity | Needs homogeneous sample | Handles heterogeneity |
| Best for | Drug target structures | Cellular architecture |
Applications
1. Cellular Architecture
Visualize organelles, cytoskeleton, and membrane systems in 3D.
2. Virus-Host Interactions
See how viruses dock, fuse, and replicate inside host cells.
3. Drug Mechanism
Observe how drugs alter cellular morphology or protein localization.
4. In-Situ Protein Complexes
Study protein complexes that are difficult to purify, such as ribosomes, proteasomes, and nuclear pore complexes.
Shuimu's TOMO Capabilities
Shuimu Biosciences offers a complete cryo-ET pipeline:
Learn more about our TOMO service or contact us to discuss your in-situ structural biology project.