I needed a detailed methodology for SEM-EDS analysis for Leaf and pollen!
I manage an SEM lab and tell researchers that they have to specify much more than "I want to do SEM on my sample". SEM and EDS can be applied in a wide variety of ways. You have to first specify what you want to know: size of features, shape of features, identity of features, number of features, distribution of features, etc.
I recommend qualitative work to begin with. Then there may reason to move to a quantitative evaluation.
You will need to say what the particulate matter is. Pollen will not be very interesting by EDS. Other particulate might be interesting.
Thank you Dr. Straszheim for replying! I want to quantify the dust particles on plant leaves exposed to ambient air based on their size (microns). Furthermore, I would like to find the elemental composition of the same particles. how should I prepare the leaf samples for analysis?
Dear Nitin Joshi ,
for the preparation task you should have a look for example at:
https://www.oxinst.com/blogs/preparation-of-biological-specimens-for-eds-with-an-sem
Best regards
G.M.
I defer to Gerhard Martens on that. White papers from instrument vendors are a good source of information. They may not be peer reviewed, per se, but you have a lot of experts in their fields putting forth their best information.
I know you will have to deal with changes in the biological material due to its water content. You will need to replace the water somehow.
If you are looking at dust particles, you probably want to use backscattered electron imaging so the dust particles stand out clearly from their organic background. You might be able to use feature analysis to automate the analysis process. Be careful of the statistics.
That was a career ago when I was doing automated image analysis of mineral matter in coal. You can look up my dissertation and papers.
It is necessary to understand that EDS fundamentally cannot replace a normal chemical analysis, especially in biological samples. In general, it is a pretty "coarse" method with lots of inescapable artifacts, which is suitable only for qualitative analysis and mapping, but not for real quantitative studies. Yes, using etalons and advanced sample preparation, sometimes it is possible to get some reliable semi-quantitative results for hard samples, especially with heavy elements, but not for biological stuff.
I'd recommend a more traditional way - just wash your leaves, sediment the particles via centrifugation and analyze the residue chemically. You can visualize them with an SEM too, using some standard preparation protocols for particles. Trying to visualize the dust directly on the leaf surface, you'll have a problem - the protocols of sample preparation for SEM typically include some kind of chemical fixation and dehydration, which obviously are wet procedures and the particles likely will be just washed out from the surface...
Good luck!