Volume Electron Microscopy with 64 Beams and Optical Transmission Detection

Imaging across multiple scales can provide valuable insights into complex biological systems, thereby enhancing the understanding of physiology in healthy and diseased states. Electron microscopy (EM) is a technique that resolves the nanoscale structure of tissues and cells on millimeter length scales, thus making it an effective tool for studying intricate biological processes. Recently, several EM techniques have been established that reveal the three-dimensional structure, collectively referred to as volume electron microscopy (volume EM).

Traditionally, 3D reconstructions of tissue and cells are achieved by cutting serial thin sections of resin-embedded samples, mounting them on support grids, and imaging with transmission EM. Today, volume EM includes several complementary techniques, each with different resolutions and field-of-view. For example, in array tomography, serial sections are placed on a solid substrate and imaged with scanning EM. In serial block-face scanning EM, a thin tissue slice is removed by an in situ ultramicrotome, and the exposed tissue block face is imaged. With a focused ion beam in a scanning EM, an even thinner slice can be precisely removed. The expanded toolkit has extended volume EM beyond its original application in neuroscience to a wide range of fields.

Advances in volume EM have largely been made possible by improvements in instrumentation, such as more automated workflows and faster and sensitive detectors. Nevertheless, the limited throughput of EMs remains a major bottleneck, especially for large volume imaging. Recent methodological innovations are, however, making possible the imaging of millimeter-sized samples and small organisms. In transmission EM, the throughput is limited by time-consuming sample grid replacement, stage movements and limited fieldof- view at high magnification. Reel translation systems with transparent tape, faster sample stages, larger camera arrays and advanced beam deflection have solved these bottlenecks and increased throughput…