Electron Tomography

Electron tomorgraphy (ET) is a technique that retrieves three-dimensional (3D) structural information from a tilt series of two-dimensional (2D) projections that is imaged at several different angles. This method provides nanometer resolution from thick samples (100 nm to 1 mm) to provide more information about the depth of the sample.

While the electron beam is stationary, the specimen holder can rotate and the specimen can be tilted at 1 – 2° angular increments over a range of ± 70 degree about a fixed axis. To prevent radiation damage to the sample, the cumulative electron dose must be kept within tolerable limits.

Equipment and Software:

FEI Tecnai G2 Twin TEM – to collect a tilt series of 2D projections

Inspect 3D software – to reconstruct 3D volume

Amira software – to align the tilt series and perform modeling

Sample Preparation:

Regular TEM sample preparation

Section thickness is in the range of 100 nm – 1 mm and thicker than normal sections

Challenges:

Optimizing Data Collection

– Staining conditions

– Pre-irradiation needed (sections may shrink due to the beam intensity)

Image Data Set Alignment

– Calculation of alignment shifts

– Adjustment of tilt axis

Publications:

Baldelli, A., Trivanovic, U., and Rogak, SK. (2019) Electron tomography of soot for validation of 2-D image processing and observation of new structural features. Aerosol Science and Technology, 1-8.

Lyon, K., Adams, A., Piva, M., Asghari, P., Moore, ED., and Vogl, AW. (2017) Ca2+ signaling machinery is present at intercellular junctions and structures associated with junction turnover in rat Sertoli cells. Biology of Reproduction 96 (6): 1288-1302.

Perez-Sancho, J., Tilsner, J., Samuels, AL., Botella, MA., Bayer, EM., and Rosado A. (2016) Stitching organelles: Organization and function of specialized membrane contact sites in plants. Trends in Cell Biology 26 (9): 705-717.