How Scanning Electron Microscopy Works
Differences Between Transmission and Scanning Electron Microscopes
Sep 20, 2009
Light microscopy can measure nothing smaller than the shortest wavelength of light because the light won't hit smaller particles. If light doesn't hit something, then nothing is visible to the human eye. So, to view extremely small objects, light microscopy is impossible to use. Electron microscopy provides the solution.
The Theory Behind Electron Microscopy
An electron has a much smaller wavelength than a photon and therefore can be used to hit and bounce off very small objects. By measuring the pattern at which the electrons are angled off their original path, one can ascertain enough about a sample that a detector catching the electrons can read out the signal in the form of a rasterized image on a screen.
Sample Preparation
Especially for biologists but for other scientists as well, sample preparation for electron microscopy is one of the most important aspects of the method. A scientist must ensure that the structures of interest are attached to or stained with a metal that is heavy enough to deflect the path of electrons. If the metal is too light, the signal is too weak and a clear image cannot be seen.
Transmission Electron Microscopy
In transmission electron microscopy (TEM) a sample is cut very thin to allow the transmission of electrons through it. Once placed on a grid with holes to allow for the passage of the electron beam, the thin slice is then placed in an electron microscope chamber where the pressure is pumped very low to allow for electrons to pass to the sample and through it without being deflected by air molecules.
Scanning Electron Microscopy
In Scanning Electron Microscopy (SEM) the sample is not cut at all but rather prepared in a whole block and placed in the chamber. Rather than have the electron beam pass through the sample onto a detector, the electron beam is shot at the face of the sample at an angle which is measured by the detector.
Scanning Versus Transmission Electron Microscopy
While it has been a common complaint of electron microscopists that SEM doesn't give as "clean" looking images as TEM, SEM will give information about the three-dimensional structure of a surface and allow for stacks of images, both of which are impossible with TEM.
TEM tends to give better signals than SEM because resolution can be higher with less interference from deeply scattered electrons that one finds in SEM samples. SEM, on the other hand, allows for imaging in the z dimension as well as in the x and y, such that one can acquire a stack of images.
With the advent of Serial Block-Face Scanning Electron Microscopy (SBF-SEM), SEM has gained popularity, though TEM still seems to predominate the field.
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