Electron Microscope Tomography and Single-particle Reconstruction

The 3D structure of biological samples can be determined using two cryo-electron microscopy techniques — electron tomography and single-particle reconstruction.

In electron tomography, a sample is progressively tilted relative to the electron beam, capturing a series of 2D images from different angles. These 2D images are then stacked using computer software to generate the 3D image or tomogram of the sample.

Electron tomography maintains the in vivo structural integrity of the sample and is primarily used for 3D visualization of cells, organelles, and macromolecules in their native environment.

In single-particle reconstruction, isolated in vitro samples, such as a purified protein sample, are evenly distributed on a grid.

A 2D image of the individual protein molecules lying at random orientations is captured. Thousands of such images are then averaged and aligned to reconstruct the 3D image of the sample.