TEM is used, among other things, to image the interior of cells in thin sections , the structure of protein molecules contrasted by metal shadowing , the organization of molecules in viruses and cytoskeletal filaments prepared by the negative staining technique , and the arrangement of protein molecules in cell membranes by freeze-fracture.
Conventional scanning electron microscopy depends on the emission of secondary electrons from the surface of a specimen. Because of its great depth of focus, a scanning electron microscope is the EM analog of a stereo light microscope.
It provides detailed images of the surfaces of cells and whole organisms that are not possible by TEM. It can also be used for particle counting and size determination, and for process control.
It is termed a scanning electron microscope because the image is formed by scanning a focused electron beam onto the surface of the specimen in a raster pattern. White light has wavelengths from to nanometers nm. The average wavelength is nm which results in a theoretical limit of resolution not visibility of the light microscope in white light of about — nm.
The figure below shows two points at the limits of detection and the two individual spots can still be distinguished. The right image shows the two points so close together that the central spots overlap. The electron microscope was developed when the wavelength became the limiting factor in light microscopes. Electrons have much shorter wavelengths, enabling better resolution.
As dimensions are shrinking for materials and devices, many structures can no longer be characterized by light microscopy. For example, to determine the integrity of a nanofiber layer for filtration, as shown here, electron microscopy is required to characterize the sample. The main SEM components include:. Electrons are produced at the top of the column, accelerated down and passed through a combination of lenses and apertures to produce a focused beam of electrons which hits the surface of the sample.
The sample is mounted on a stage in the chamber area and, unless the microscope is designed to operate at low vacuums, both the column and the chamber are evacuated by a combination of pumps. Many applications require minimal sample preparation.
Modern SEMs generate data in digital formats, which are highly portable. Samples must be solid and they must fit into the microscope chamber. Maximum size in horizontal dimensions is usually on the order of 10 cm, vertical dimensions are generally much more limited and rarely exceed 40 mm.
For most instruments samples must be stable in a vacuum on the order of 10 -5 - 10 -6 torr. Samples likely to outgas at low pressures rocks saturated with hydrocarbons, "wet" samples such as coal, organic materials or swelling clays, and samples likely to decrepitate at low pressure are unsuitable for examination in conventional SEM's.
However, "low vacuum" and "environmental" SEMs also exist, and many of these types of samples can be successfully examined in these specialized instruments. Most SEMs use a solid state x-ray detector EDS , and while these detectors are very fast and easy to utilize, they have relatively poor energy resolution and sensitivity to elements present in low abundances when compared to wavelength dispersive x-ray detectors WDS on most electron probe microanalyzers EPMA.
An electrically conductive coating must be applied to electrically insulating samples for study in conventional SEM's, unless the instrument is capable of operation in a low vacuum mode. Sample preparation can be minimal or elaborate for SEM analysis, depending on the nature of the samples and the data required. Minimal preparation includes acquisition of a sample that will fit into the SEM chamber and some accommodation to prevent charge build-up on electrically insulating samples.
Most electrically insulating samples are coated with a thin layer of conducting material, commonly carbon, gold, or some other metal or alloy. The choice of material for conductive coatings depends on the data to be acquired: carbon is most desirable if elemental analysis is a priority, while metal coatings are most effective for high resolution electron imaging applications.
Alternatively, an electrically insulating sample can be examined without a conductive coating in an instrument capable of "low vacuum" operation. Goldstein, J. Reimer, L. Springer, p. Egerton, R. Springer,
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