Embedded Files
Prepare the samples for SEM
Prepare the samples for SEM
The plastic samples need to be dehydrated, therefore water is replaced by ethanol through a series of steps in solutions with increasing concentration of ethanol.
The samples are then coated with a very thin layer of gold (Au) through a process called sputtering. This will allow the SEM to recognize the substrate.
Taking pictures with SEM
Taking pictures with SEM
To introduce a sample inside the SEM a particular sequence is required in order to obtain the vacuum inside the chamber. After the sample has been scanned by the microscope 6 random frame at 2 different zooms (500x and 1500x) are taken in order to subsequently analyze diatoms and bacteria.
SEM Image analysis for bacteria and diatom identification and quantification
SEM Image analysis for bacteria and diatom identification and quantification
The images obtained with the Scanning Electron Microscopy are analyzed using the software ImageJ, which allows to count objects with a click. Three different measurements are performed: coverage index and quantification of diatoms with the images at 500x, while quantification of bacteria is performed at 1500x.
Assessing coverage index
Assessing coverage index
The coverage index is a visual estimation that describes the level of colonization of a plastic piece. The Plastisphere is composed by several organisms and their excretions. We can find them dead or alive, isolated, in colonies or as biofilm.
The index ranges from 0 to 4 where:
0=0% absence of OM
1=25% OM coverage
2=50% OM coverage
3=75% OM coverage
4=100% no plastic can be been, fully covered.
0.5 decimals can be used as well.
Identification and counting of diatoms
Identification and counting of diatoms
Diatoms are among the main colonizers of MPs. The characterization and counting of diatoms is performed at a magnification of 500x. It generally allow us to obtain a classification at the genus level. Some examples of what I found in my samples are in the figures.
Identification and counting of bacteria
Identification and counting of bacteria
We distinguished bacteria morphologies on images with a magnification of 1500x. We classified them according to their shape:
Cocci (in yellow) - round shape
Bacilli (in pink) - elongated shape
Bacillococci (in blue) - a mix of the previous ones
Bacteria are often found in aggregates or covered by biofilm.
Guess how many bacteria are in this image...
Guess how many bacteria are in this image...
answer=494!
Page updated
Google Sites
Report abuse