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Nick S
Lv 5
Nick S 發問於 Science & MathematicsBiology · 1 十年前

E. coli bonding to a surface?

I have a butyr aldehyde treated thermal oxide (SiO2) surface on a silicon wafer, and am trying to figure out a way to get E. coli to stick to the surface. I have thought of how to keep them alive. When incubating the disc, I just need to keep them in a minimal growth medium. When washing with PBST, I should likely actually mix PBST and the minimal growth medium to keep them from going dormant during the wash.

To bond them to the surface, would a simple antibody print on the disc work? I am considering trying to use a polyclonal anti-rabbit antibody from Abcam, but am unsure if that will be enough to grab onto the bacterium (the antibody is roughly 1/1000 the size of the bacterium), and don't want to make a hasty purchase if it definitely won't work ($300/mg of antibody).

Is there another direction I should be looking to go? Poly-lysine is not an option, unfortunately, as I am trying to observe a phenomenon that is very height sensitive...

Any ideas?

更新:

swd:

Thanks for trying. I know this one isn't a very easy question...I've been trying to think of it for a while now, and every time I think I have my finger on the answer, something comes to mind, and it becomes a little less clear what to do...

更新 2:

They are treating the surface with butyr aldehyde, and the first attempt is apparently going to be a slow flow of E. coli suspended in growth medium, to see if any will bond directly to the surface. It seems like the antibodies have been put off for now, but I believe that there is a company that will print antibody spots on the thermal oxide layer for me when the time comes. What I'm not sure of is how effective this bonding (either with butyr aldehyde or the antibody) will be. Also, we will not need the conjugated antibody, so it's just the goat polyclonal IgG in question.

And while we do not necessarily need a single layer, the phenomenon I am hoping to observe would be most obvious in a single layer (even a few isolated bacteria on the plate could yield the desired result).

The idea is to hit them with an antibiotic and observe a protein bloom in real-time as the cell membrane degrades, but the first challenge is to get them to stick to the surface at all.

Thanks

1 個解答

相關度
  • H37Rv
    Lv 4
    1 十年前
    最愛解答

    Do you need bacteria in a single layer? If not, would it be possible to induce a biofilm formation and get the bacteria to adhere themselves to the surface?

    The antibody should be enough to hold onto the bacteria because not one but many antibodies will grab the bacterial surface, the question is how do you intend to get the antibody to stick to your surface? Antibodies stick to poly styrene really well, any protein does, but I don't know anything about sticking to SiO2. Can you conjugate it to something that will stick?

    Edit: I would look into what stimulates biofilm formation in E. coli, this might be a good way to get them to attach to the surface. If you find the right conditions it will increase the chaces that the bacteria will be expressing the necessary adhesins for attaching to a surface.

    http://jb.asm.org/cgi/content/full/188/2/587

    This paper says that biofilms are inhibited by Autoinducer-2, the quorum sensing molecule. So I would recommend using an e coli culture that is not dense, i.e. early log phase.

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