Bay tree pollen
Determining the optimal working conditions for your electron microscopy experiment is vital as is understanding why certain variables should be considered, and the team at the  Plymouth Electron Microscopy Centre (PEMC) will help you set up your experiment with your particular goals in mind. 
This short case study gives a visual example of how the accelerating voltage of the instrument used (and the subsequent interaction volume) has key implications on both imaging and compositional analysis.
Interaction Volume
The interaction volume of the electron beam is something that you'll hear electron microscopists talking about a lot. This is the volume of the sample that is affected by the electron beam during analysis. Higher accelerating voltages produce larger interaction volumes, and lower accelerating voltages produce smaller interaction volumes. As such, if you are interested in imaging small features then a lower accelerating voltage is most suitable, provided enough signal is generated for the detectors being used. If the voltage is too high, then the finer details will be lost due to the increased interaction volume. 
It can often be difficult for users to picture the interaction volume in relation to their sample, as it isn't immediately obvious from the images generated. Software such as Casino (Monte Carlo Simulation of Electron Trajectory in Solids) generates useful graphs of the interaction volume at different accelerating voltages and in different materials. 
The graphs below were generated by Casino. These reflect the interaction volumes at 5 keV, 10 keV, and 20 keV in a sample of pure iron (1 keV = 1000 V). As is clearly shown here, the higher voltages produce significantly larger interaction volumes. 
How interaction volume impacts imaging
As mentioned above, lower accelerating voltages (and therefore smaller interaction volumes) allows for the imaging of finer details than higher voltages. This is a secondary electron image of bay tree pollen taken on our JEOL IT800 FE SEM . The left side of the image was taken at 5 keV, and the right at 20 keV. 
Not only did the higher voltage obscure some of the detail on the pollen surface, it also caused obvious sample damage. During imaging, the pollen was observed to blister and crack in real time. Sample damage is particularly important for beam sensitive and biological samples, and lower accelerating voltages are typically used to avoid this, as well as increasing resolution.
Bay tree pollen
Voltage difference
For some sciences, samples are prepared as polished blocks for high quality compositional analysis. This sort of analysis typically utilises 15-20 keV which can hide a lot of the surface defects on the sample. This is an example of a meteorite that has been polished for Energy Dispersive Spectroscopy (EDS) analysis. The left side of the image was taken at 20 keV and the right at 3 keV. 
Here, it is very apparent that there are many fine scratches and surface contamination that can be observed at low acceleration voltages and although minor, these features can have an effect on the quality of data produced. 
We can also see a blurring of crystal boundaries at higher accelerating voltages due to the larger interaction volumes, and some examples of this are below. 
Balancing interaction volume and overvoltage
Overvoltage refers to the voltage required to sufficiently excite elements during compositional analysis to be able to measure them, and is typically twice as much as the energy of the element of interest. For instance, the iron K-alpha line has an energy of 6.4 keV, so a voltage of almost 13 keV would, traditionally, be ideal to quantify it. Developments in detectors allow for low energy analysisr  and reduced overvoltages, which means that compositional analysis can be conducted at lower accelerating voltages and improved spatial resolution than previously possible. 
However, it is still important to ensure that the energies used are appropriate for the elements in the sample and this comes with the caveat of larger interaction volumes. It can therefore be beneficial to combine lower voltage imaging with higher voltage EDS analysis to fully understand a sample. The software available at PEMC allows for some degree of automation, meaning that a series of images could be collected under one set of working conditions and then compositional analysis at the same points under a different set of working conditions. 
If low energy compositional analysis is useful for your experiment, please get in touch with the lab to discuss the instrumentation and options available to you.