Dear Dr. Barkovskii, strong biofilm formation on the surfaces depends on surface properties of material surface, as well as surface properties of bacteria. In our previous works, we isolated some S. epidermidis strains from biomaterial surfaces which are polymeric materials. They strongly adhere and form biofilm and we cannot remove from surfaces by washing. Additionaly same S. epidermidis strains strongly adhere stainless steel surfaces. 

Sincerely yours.

It depends on the strain(s) that you are trying to make a biofilm with. I usually find which strains are the strongest biofilm former's first. I   use a range of temperatures from 10 C to 30+ C to enhance biofilm formation and also incubation times (1 day to 9 days). Sometimes I add 2% glucose/ alcohol/salt to the medium to  promote biofilm formation. To date the best biofilms I have made are from mixed species environmental isolates.

 Thanks Mert and Gerry,

I have also characterized the strains first, and only one of these, P aeruginosa and not so spectacular biofilm producer formed a strong biofilm.Mert, can you send me that S. epidermidis? Gerry can you guide me how do you make biofilms?  I also tried to mix environmental isolates with no strong outcome. Cheers,

Dear Andrei, I will check the strains. I'm now outside of the my lab. I will give you about availability of strain.

Best regards.

Thanks, Mert!

Here are some steps I have taken to ensure good biofilm.

In my work with Staphylococcus, not all species formed biofilms under the set of conditions I describe but i could stress the organisms with different parameters and most would form biofilms.

Usually I would start by growing the organism overnight so that it is well into stationary phase. The next morning I would make a 1/100 dilution of the culture and dilute it with fresh growth medium . Typically this medium would be 1/2 --> 1/5 th strength media . Many of my experiments were then conducted on 96 well high bind plates . High bind is important for good biofilms since it is compatible with many proteins . Low quality highly charged plastic  has never been that good in my hands. I also grew a lot of my biofilms at  room temperature not 37 C. My biofilms would take about 3 days to mature. I would cover the 96 well plate with foil and usually keep it in a dark place , ensuring the the surface it was on was completely level. For environmental biofilms, i cultured these at a slightly lower temperature for a maximum of 9 days depending on the source. Some species I found it extremely difficult to cultivate into biofilms but sometimes the addition of 2% glucose or alcohol will stress the organism out enough to form a biofilm. You can also try coating the surface of your plates with dilute solutions of single or multiple amino acids or poly lysine. however these modifications also make the interpretation of any biofilm changes complicated.

If I have missed anything , the rest will be in https://www.ncbi.nlm.nih.gov/pubmed/25359476. 

Thanks,Gerry.  Will you please send me a copy of the above manuscript.

Yes Andrei, I just posted the manuscript.

If you are working on environmental biofilms these are a lot harder to cultivate and require slightly more time to grow.  I should be bringing a paper out soon on marine biofilms.

Thanks, Gerry, I got the manuscript. Yes, in this project I am working on environmental/industrial biofilms or rather on their reliable removal from waterlines, pipelines, and alike.

Cheers, 

Ha Gerry

the amino acid approach worked, thanks for the suggestion.

Cheers,