There are planty ways to do that. if you already have done biofilms in 96 well plates, you can do in the same plates, grown the biofilm , remove plantonic cells, and then collet biofilm by pipeting with PBS and perform CFU count in agar.
There is a special Deviace similar to 96 well plates, where you can calculate MBEC (Minimal biofilm erradication concentration) Here is the reference.
"The MBEC Assay System: multiple equivalent biofilms for antibiotic and biocide susceptibility testing.
Ceri H1, Olson M, Morck D, Storey D, Read R, Buret A, Olson B."
Also i have seen some report with microfermentator, but i dont now much about it.
Alternative technique by fluorescent study by taking some agents/quenching chemicals
Dear Dinesh, you check biofilm formation on material surfaces by simple staining with crystal violet then visualize optic microscope or stereo zoom microscope. If it is available, you can use SEM and AFM techniques for detailed analysis
If you want to quantify biofilm cells on material surfaces, first you can remove unattached cells then detach biofilm formers by cell scapers and sonication. It depends on your material.
Best regards.
but how to grow pathogens or develop biofilm ? on which material or surface I should grow it ? I want to check my nanomaterial antibiofilm activitity.
I do not know a standardized method but you can wash the surface with dist water and dry at 40-50C up to a constant value and weigh the sample and compare with the control.
Hi Dinesh,
One of the easiest method is crystal violet staining. You can use polystyrene microtiter plate. For your reference, please check the articles the links of which are mentioned below. I hope that would be helpful to you.
https://www.nature.com/articles/srep44419.pdf
https://www.nature.com/articles/srep18877
Devapriya
Hi Dinesh,
If you have different types of 96-well plates available you can check and see what type of material the biofilms will attach and grow the best. Some bacteria work better on polyvinyl chloride (PVC) and others on polystyrene. Using the crystal violet method is probably the easiest way to quantify the biofilm inhibition of your nanomaterial.
Also check the supplementation of the media with iron, as in some cases the addition can increase the biofilm formation.
Kind regards
Dear Dinesh,
First and foremost things is to understand which study you want to do: Biofilm inhibition study or biofilm disruption. If you want to do biofilm inhibition, you will have to incubate your compounds and the bacteria together. Mind you, the concentration of the compound has to be at least 1/4 MIC for inhibition. You can grow them on 96 well plates or even better on cover slips in 12 well or 6 well plates (do not shake the plates while they are in the incubator). Depending upon the bacteria, media and conditions can be established. Removal of the planktonic cells and washing the coverslip with PBS to remove all planktonic cells is important before you quantify or stain the biofilm. For conducting disruption study, you will have to first grow the bacteria into a biofilm. This can be done by the steps mentioned above, only difference being, there is no addition of compound. After the biofilm has formed (at least 24h). You can wash off the planktonic cells, incubate the biofilm with compounds of interest at different concentrations. After 24h, you check how much of the biofilm has been disrupted using the same procedure as mentioned in the previous posts.
Hope it helps.
CD
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