I want to do some work about biofilm now. However I failed in getting a stable biofilm out of S.aureus . I followed a method that said "S. aureus (5 × 107 CFU/ml) was grown in Luria-Bertani (LB) broth on 96-well plates at 37◦C for 24 h”. But when I removed the lanktonic bacteria from the 96 well plate,it seemed that there was nothing left(no obvious film), all the cells sitting at the bottom were removed togther with LB, not to mention after washing twice with PBS. Then I tried by prolonging the incubation time to 2 days even 3 days, but still no film formed.
I hope someone who have the experience to grow biofilm can help me out. thanks!
S. aureus will form stronger biofilm in the culture containing 1% - 2% glucose. You can try biofilm assay with both glucose (-) and glucose (+) to compare the results. Besides, you can try with the broth TSB (trypticase soy broth) instead of LB.
As Mi said, you can use different media. After that if you do not see biofilm, it means that your strain does not form biofilm.
Did you use Crystal Violet for visibility of biofilm?
Fang Ji Yes, testing different strains at the same time is the best choice without losing time. Beside this I suggest you to use Crystal Violet once, even when your biofilm is not stable because some times you have biofilm but it is not visible without staining.
I highly recommend TSB. How many strains of SA did you examine? The non-forming biofilm strain of SA is unusual, so I think the medium may be the problem, and the best option would be to repeat the experiment with LB and TSB as a comparison. Also, pay attention to the technique you use to remove planktonic cells because the tip can easily scratch the biofilm off. And consider omitting rinsing with PBS because some parts of the biofilm will be detached.
Please include at least one type strain ATCC, 6538 and 33591 form biofilm for sure :)
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