Important Factors to Consider for Electron Microscopy

Electron microscopy has increasingly found more applications in recent years. Every sample has a mixture of optimized settings that must be utilized to enhance the results of the investigation.

This article will outline all of the key aspects that need to be taken into account when imaging samples and will outline some of the mathematics and physics behind them.

1.1 Magnification

Magnifying glasses were first found in Ancient Greece, where Aristophanes outlined the first attempt to view closer details as a children’s leisure activity. This was when the term ‘magnification’ first entered human language.

The interest in science for the nano and micro world has greatly increased as time has passed, opening up the requirement for a quantification of magnification.

Magnification is defined in modern times as the ratio between two measurements, which suggests that two objects are required for an accurate assessment of the value.

The sample is obviously the first object and the second is an image of it. While the sample will not vary in size, the image can be printed in an infinite number of various sizes.

This means that printing an image of an apple that is the correct size for a regular printer sheet and printing it again to fit on a poster that will be utilized to cover a building, will significantly adjust the magnification value (much bigger in the second example).

An example that is more scientific can be found in the context of microscopy: when storing a digital image of the sample, changing the size of the image results in the magnification number becoming clearly incorrect.

Magnification is a relative value which means it is not helpful in the field of science.

Scientists utilize two parameters that outline the specific imaged area. These are the field of view which is the region that the microscope points at and how sharp the image is, known as the resolution. The formula of magnification also varies accordingly:

The formula still clearly provides a brief description and does not factor in the resolution. This means that the magnification number will change when scaling the same image to a larger screen.

Image Credit: Thermo Fisher Scientific Phenom-World BV

The size of the object to be imaged is defined by the field of view. This value normally varies between several millimeters (a bug) to several microns (the hair of a bug) and a couple of nanometers (the exoskeleton’s molecular macrostructure).

Objects in the range of several hundred picometers can be imaged using modern tools which is the standard size of an atom. How does one define the necessary field of view to image a sample? It depends.

As an example, if the particles have an average size of 1 micron and the application requires them to be counted, it is sufficient to have 20 particles per image, instead of wasting time by imaging one particle at a time.

Even when the empty space between particles is taken into account, a field of view of 25 to 30 microns is adequate for such a sample. If the particle structure is the focus of the investigation, a close up is necessary and the recorded area must be nearer to 2 to 3 microns, if not less.

Images of particles. A close-up of a particle (left) shows the surface topography (FOV=92.7 µm). A larger field of view (right) enables more particles to be imaged (FOV=µm).

Images of particles. A close-up of a particle (left) shows the surface topography (FOV=92.7 μm). A larger field of view (right) enables more particles to be imaged (FOV=μm). Image Credit: Thermo Fisher Scientific Phenom-World BV

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This information has been sourced, reviewed and adapted from materials provided by Thermo Fisher Scientific Phenom-World BV.

For more information on this source, please visit Thermo Fisher Scientific Phenom-World BV.

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