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Volume EM with Multibeam Array Tomography
Volume EM Techniques

Multibeam Array Tomography​

High Throughput of Large Volumes of Ultrastructural Data​
  • Very fast acquisition of ultrastructural details
  • Sample sizes that cannot be imaged with other technologies
  • Statistical characteristics with previously unachievable quantitative significance

Volume EM with Multibeam Array Tomography

In array tomography (AT), serial sections are imaged by SEM and then digitally reconstructed to create a 3D data set. A multibeam SEM (ZEISS MultiSEM) combines up to 91 electron beams to significantly accelerate image acquisition. Multibeam AT can image volumes larger than a cubic millimeter at nanometer resolution, ideal for fields such as connectomics. Multibeam AT could even be used for mapping of large neuronal networks, such as the entire mouse brain.​

Schematic Representation of a Typical Workflow

Multibeam Array Tomography sample preparation

1

A resin-embedded sample is cut into an array of serial sections, each with a section thickness of typically 30 – 70 nm, and attached to a sample carrier in the order they were cut.​

Multibeam Array Tomography image acquisition

2

Each serial section is imaged in the multi-beam scanning electron microscope (ZEISS MultiSEM).

Processing segmentation

3

The acquired EM images are processed and digitally aligned into a 3D data set. Cell compartments can be identified and segmented. ​

3D visualization analysis

4

The segmented 3D data set can be visualized, investigated, and statistically analyzed. ​​

Application Examples​

Understanding Neuronal Connectivity of Brain Tissue on a Larger Scale

Statistical Characteristics of Neuronal Connections with Previously Unachievable Quantitative Significance

The brain is a complex organ with millions of neuronal connections and signaling pathways. Understanding the relationship between the structure and function of brain tissue will help to unravel complex neuronal networks and, in the long term, how to treat certain pathologies with medical interventions.​

​Large-scale, high-resolution imaging helps to explore these neuronal connections in the brain. For small brain specimens, the time required to capture a full 3D data set is significant, but achievable with the right technology. By utilizing multiple electron beams in parallel, ZEISS MultiSEM enables the acquisition of image data with unprecedented speed, making imaging of an entire brain from a mouse possible.​

​The images below show various zoom factors of the same sections from a mouse brain imaged with ZEISS MultiSEM with 61 electron beams. Sample courtesy of J. Lichtman, Harvard University, USA.

Assembled mosaic of a square millimeter captured at 4 nm pixel size in 6.5 minutes from a 30 nm thick brain slice, prepared with a high-contrast staining protocol and cut with an ATUMtome, an ultramicrotome that collects sections on a tape.
Assembled mosaic of a square millimeter captured at 4 nm pixel size in 6.5 minutes from a 30 nm thick brain slice, prepared with a high-contrast staining protocol and cut with an ATUMtome, an ultramicrotome that collects sections on a tape.

Assembled mosaic of a square millimeter captured at 4 nm pixel size in 6.5 minutes from a 30 nm thick brain slice, prepared with a high-contrast staining protocol and cut with an ATUMtome, an ultramicrotome that collects sections on a tape.

Individual hexagonal multibeam fields of view (mFoV) put together using an exemplary set of seven mFoV taken from the previous data set.
Individual hexagonal multibeam fields of view (mFoV) put together using an exemplary set of seven mFoV taken from the previous data set.

Individual hexagonal multibeam fields of view (mFoV) put together using an exemplary set of seven mFoV taken from the previous data set.

Individual hexagonal multibeam fields of view (mFoV) put together using an exemplary set of seven mFoV taken from the previous data set.

Example of a single mFoV, consisting of 61 individual image tiles acquired with 61 electron beams in parallel, covering more than 100 µm from left to right, typically acquired in just seconds.
Example of a single mFoV, consisting of 61 individual image tiles acquired with 61 electron beams in parallel, covering more than 100 µm from left to right, typically acquired in just seconds.

Example of a single mFoV, consisting of 61 individual image tiles acquired with 61 electron beams in parallel, covering more than 100 µm from left to right, typically acquired in just seconds.

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