I would like to test a chemical to determine its activity on the biofilm matrix. I already know that it reduces biofilm formation, but it is not microbicidal. I want to separate the biofilm matrix (EPS and other substances) from microbial cells in several monospecies biofilms to see if the chemical interferes with matrix molecules using EPR. Can anyone suggest a good and proved protocol to separate matrix from cells?
Thanks
I don't have much experience with biofilms in particular, but maybe something along the lines of density gradient centrifugation with HistoDenz may help?
Dear Elisa, Sometime ago I was trying to isolate the EPS from our Pseudomonas fluorescens SBW25 biofilm, and we found that submerging the biofilms in distilled water over-night removed up to 90% of the viable cells - presumably they all moved up the water column following the oxygen gradient. After several over-night incubations we had a substantially cleaner matrix that we subjected to nucleases, lipase and proteases to purify the EPS. If you have the time and your biofilms are not too delicate, simple procedures like this can be useful. Regards, Andrew.
This is interesting. I don't need to use enzymes to digest everything but EPS, because I want to get the entire matrix. I could compare the methods and see which one gives a higher matrix amount. Which way would you suggest to quantify the matrix after separation?
Hi Elisa,
Although I haven't used the technique myself but from what I have read, the use of cation exchange resins can extract the matrix components without compromising cell intergrity. Of course you will have to optimise the assay to work with your samples. The attached paper might be a good place to start. All the best.
Esther.
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