Using SEM Technology in Electron Microscopy Analysis

There are a number of essential parameters which must be accounted for when undertaking electron microscopy (EM). When correctly taken into account, these can help produce the best possible results and ensure that the sample’s features are imaged appropriately.

Voltage, or tension, plays a critical role when applied to source electrons to generate an electron beam. Generally, the trend has always been to increase voltage in order to improve the system’s resolution.

In recent years however, producers of scanning electron microscopes (SEM) have begun to focus on improving resolution at lower voltages. This has been largely prompted by the increasing use of EM within life sciences, particularly following the introduction of the Nobel prize-winning cryo-SEM technique.

Electron Beam Voltage Shapes the Interaction Volume

Electrons’ energy content is indicated by voltage, so this will determine how a beam will interact with a sample. Typically, an increase in voltage will result in an increase in penetration below the sample’s surface. This amount of penetration is known as interaction volume.

When this occurs, electrons will have larger, deeper propagation within the sample, thus generating signals within different parts of the affected volume. A sample’s chemical composition can also affect the size of the interaction – for example, light elements have fewer shells, so the electrons’ energy content is lower. This then limits interactions with electrons from the electron beam, meaning it can penetrate deeper into the sample than it would be able to with a heavier element.

Different results can be acquired when analyzing outcoming signals. Within desktop instruments, there are three kinds of signals that are usually detected – X-Rays, backscattered electrons (BSE) and secondary electrons (SE).

Effects of Voltage in SEM Imaging

Voltage effects BSE and SE imaging in comparable ways, with low voltages enabling the surfaces of samples to be imaged while higher voltages can provide more information on layers beneath the sample’s surface.

The images below illustrate this with practical examples, where low voltages clearly highlight surface sample contamination while increased tensions reveal structures on the surface below the contamination layer.

BSE images of tin balls at 5 kV (left) and at 15 kV (right). With the lower voltage, the carbon contamination on top of the sample becomes visible. When the voltage is increased, the deeper penetration enables the imaging of the tin ball surface underneath the carbon spots.

Figure 1. BSE images of tin balls at 5 kV (left) and at 15 kV (right). With the lower voltage, the carbon contamination on top of the sample becomes visible. When the voltage is increased, the deeper penetration enables the imaging of the tin ball surface underneath the carbon spots.

The sample’s nature is also a defining factor when choosing an appropriate voltage. Several polymers, biological samples and a range of other (predominantly organic) samples are highly sensitive to the high energy content of electrons. This sensitivity becomes more prevalent as SEM operates within a vacuum.

Due to this, SEM developers are continually striving to increase resolution at lower voltages, thus providing results with even the most delicate of samples.

The key problem with this process is the imaging technique’s underlying physics principle. In a comparable way to photography, there is a range of distortions and aberrations that can affect the quality of the final output.

When using higher voltages, chromatic aberrations become less of an issue, and this is the core reason that SEM has historically tended to use the highest possible voltage in order to improve imaging resolution.

Generation of X-Rays in a SEM

With X-Ray generation however, the story is a wholly different one. High voltage results in a higher production of X-Rays, and these can be captured and then processed by an energy dispersive spectroscopy (EDS) detector in order to perform compositional analysis on a sample.

This technique involves forcing the ejection of an electron within the target sample by interacting with electrons (primary electrons) from the electron beam.

When this occurs, a charge vacancy – effectively, a hole – is generated within the inner shells of an atom and this is then filled with an electron with a higher energy content that was originally in an outer shell of the same atom.

For this process to work it requires the electron to release part of its energy in the form of an X-Ray, and this X-Ray’s energy can then be correlated to the atomic weight of the atom via Moseley’s law, returning the sample’s composition.

The core factors in X-Ray production are:

  • Interaction volume: This defines the spatial resolution of the analysis.
  • Overvoltage: This is the ratio between the energy of the incoming beam and the energy needed to ionize the target atom.

In order to achieve the best possible analysis, it is important to reach a minimum overvoltage value of 1.5. This means that by increasing the electron beam voltage, the maximum number of elements which can be detected is increased.

In contrast, a high voltage corresponds with a high probability of sample damage and, more importantly, a larger interaction volume.

When this happens, not only is a sample’s reliability potentially compromised, but also, the generation of X-Rays interacts with a far larger volume. When working with particles, multilayers and otherwise non-isotropic materials, having a larger interaction volume will generate signals which come from parts of the sample with varying compositions, thus compromising the quality of results presented.

Example of an EDS spectrum collected at 15 kV. The peaks highlight the presence of an element and a complex algorithm is applied to convert the signal coming from the detector into chemical composition.

Figure 2. Example of an EDS spectrum collected at 15 kV. The peaks highlight the presence of an element and a complex algorithm is applied to convert the signal coming from the detector into chemical composition.

Generally, recommended tension levels for the analysis fall between 10 and 20 kV in order to balance these two effects. Selecting the ideal value very much depends on an additional element of EDS analysis known as ‘peak overlap’.

X-Rays generated by electrons moving from different shells of different elements will often have comparable energy contents, and this means that more advanced integration processes are needed in order to deconvolute the peaks and normalize the results. Alternatively, it is possible to use higher energy content lines which come from one of the two elements with overlapping peaks.

Most EDS software includes functionality that automatically applies the former of the above two options, while the latter is sometimes impossible given the higher energy level line for a common element (for example, lead) would likely require a voltage of over 100 kV.

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

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

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