How to stain biofilms in a 96-well plate

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[tartanhandbag]

you may find that the initial washing steps submerging in water create hydrodynamic aberrations strong enough to disturb and remove all or parts of your biofilm in weak biofilm forming bacteria. in such cases you may wish to pipette in the dH20 or PBS in and out of the well (still doing it 3 times). it takes a bit longer but is important if you are struggling to keep your biofilms in tact. you can monitor this throughout the biofilm assay by regularly inspecting the plate by eye to see what lies at the bottom of the wells before and after each treatment.

[aliyahahmad]

What kind of medium that you used in this assay ?

[nahidbodiz]

when you pipet up and down the biofilm with ethanol, the cells distributed in a well could create a heterogeneous solution. Can we transfer from each well to the microtube then centrifuge and read a clear solution in the spectrometer?

[BiankaRinaldinFonsecaFerreira]

Hi Lisa! Thanks for the video! It helped a lot! Just a question, some protocols uses 33% acetic acid solution instead of ethanol, do you know how can I prepare this solution? Is it water or alcohol?

[luisa]

Could you add in the description the species you are using for your biofilms test? And just in case: crystal violet is a toxic compound. In Germany it is not allowed to be poured in the sink. It has to be collected and disposed by a company. All water organisms and the water treatment microbes will be damaged if pouring chemicals down the drain. Inform yourself about disposal procedures in your country.

[dalalshowali]

Thanks alot Dr. The video is wonderful. I have one aquastion, what is the wavelength suitable for reading the optical density for a biofilm stained with crystal violet and MTT ?

[radia]

May be it's a bad question but I want to know how you prepare your CV solution?

[monikafelegyhazi]

Hello ! Can you help me ? Pektin liaz enzim can destroy biofilm ?
How can it use ?
10mg 1liter water ?

[MIrfanArif]

I am worried about contaminating other stuff. We are not allowed to drain the media in the sink at all. And if we use pipetting to wash from each cell, would it disturb the biofilm?

[kamillaanochshenko]

Could you please tell me what microorganisms were used for this experiment to grow biofilm? Also, in different protocols different wavelength is used to measure the OD of the plate, could you please explain why is that so? Is that related to what chemical is used to solubilise crystal violet?

[Veterinaryscitec]

Mam I am taking the absorbance at 620nm .. does it make any difference in the readings

[hagarsaid]

Is this microtiter plate specially prepared for Pseudomonous?

[dwifebrianti]

Do we need to air-dry plate befor adding the ethanol?

[raedamer]

Good evening doctor, how are you? Thank you for the nice video. Can you explain the steps in full, for example, how to add bacteria, and after that the blue dye and after it the ethanol. I am a major master’s student, Amer from Iraq Micro Biology

[hagarsaid]

Could you please explain the method from the start?

[lapetitebantam]

Isn't ethanol normally damaging the membrane of bacteria?

[منتدىشبابالرافدين]

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