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Preparation of Samples for Electron Microscopy01:20

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To be visualized by an electron microscope, either transmission or scanning, biological samples need to be fixed (stabilized) so the electron beam does not destroy them and dried thoroughly (desiccated/dehydrated) so the vacuum does not affect them. Fixation needs to be done as quickly as possible because the sample properties will start changing as soon as it is removed from its natural environment. For example, in a tissue sample, the oxygen levels begin decreasing, causing an altered...
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In 1931, physicist Ernst Ruska—building on the idea that magnetic fields can direct an electron beam just as lenses can direct a beam of light in an optical microscope—developed the first prototype of the electron microscope. This development led to the development of the field of electron microscopy. In the transmission electron microscope (TEM), electrons are produced by a hot tungsten element and accelerated by a potential difference in an electron gun, which gives them up to 400...
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A scanning electron microscope (SEM) is used to study the surface features of a sample by using an electron beam that scans the sample surface in a two-dimensional manner. Typically, areas between ~1 centimeter to 5 micrometers in width can be imaged. SEM can be used to image bacteria, viruses, tissues as well as larger samples like insects. Conventional SEM gives a magnification ranging from 20X to 30,000X and spatial resolution of 50 to 100 nanometers.
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Overview of Electron Microscopy01:25

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The wavelengths of visible light ultimately limit the maximum theoretical resolution of images created by light microscopes. Most light microscopes can only magnify 1000X, and a few can magnify up to 1500X. Electrons, like electromagnetic radiation, can behave like waves, but with wavelengths of 0.005 nm, they produce significantly greater resolution up to 0.05 nm as compared to 500 nm for visible light. An electron microscope (EM) can create a sharp image that is magnified up to 2,000,000X.
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Transmission electron microscopy (TEM) can be used to determine the 3D structure of biological samples with the help of techniques such as electron microscope tomography and single-particle reconstruction. While single-particle reconstruction can examine macromolecules and macromolecular complexes in vitro conditions only, tomography permits the study of cell components or small cells in vivo.
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Cryo-electron Microscopy Specimen Preparation By Means Of a Focused Ion Beam
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Using Xe Plasma FIB for High-Quality TEM Sample Preparation.

Suzy M Vitale1, Joshua D Sugar2

  • 1Earth and Planets Laboratory, Carnegie Institution for Science, 5241 Broad Branch Rd NW, Washington, DC20015, USA.

Microscopy and Microanalysis : the Official Journal of Microscopy Society of America, Microbeam Analysis Society, Microscopical Society of Canada
|March 15, 2022
PubMed
Summary
This summary is machine-generated.

High-quality transmission electron microscope (TEM) samples can be prepared using xenon (Xe) plasma focused ion beam (PFIB) milling, comparable to gallium (Ga) focused ion beam (FIB) methods. Xe PFIB offers large-area TEM sample preparation, with a trade-off between thickness and transparency area.

Keywords:
EELSFIBPFIBTEM sample preparationgalliumxenon

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Area of Science:

  • Materials Science
  • Analytical Chemistry
  • Microscopy

Background:

  • Transmission electron microscope (TEM) sample preparation is crucial for high-resolution imaging.
  • Gallium (Ga) focused ion beam (FIB) is a common technique, but alternatives are sought.
  • Xenon (Xe) plasma focused ion beam (PFIB) offers potential for larger area sample preparation.

Purpose of the Study:

  • To directly compare the quality of TEM samples prepared using Ga FIB and Xe PFIB.
  • To determine if Xe PFIB can produce equivalent quality samples to Ga FIB.
  • To establish best practices for Xe PFIB TEM sample preparation.

Main Methods:

  • Samples were prepared using both Ga FIB and Xe PFIB.
  • Deposition and milling parameters were varied during sample preparation.
  • Sample thickness was evaluated using scanning TEM-electron energy loss spectroscopy (STEM-EELS) t/λ data.

Main Results:

  • High-quality, large-area TEM samples can be prepared using Xe PFIB.
  • Xe PFIB sample preparation is feasible and comparable to Ga FIB.
  • A trade-off exists in Xe PFIB between ultimate sample thickness and electron transparent region size.

Conclusions:

  • Xe PFIB is a viable alternative to Ga FIB for preparing high-quality TEM samples.
  • Best practices for Xe PFIB workflow enable consistent success.
  • Optimization is needed to balance sample thickness and transparent area in Xe PFIB preparation.