Array antibodies onto membranes - (Aug/24/2006 )

Hi,

Anyone familiar with spotting antibodies onto membranes? Which brand of membrane should I use and how much antibodies to use?

thanks

Muly

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Please more details. Which substrate are you doing with and what your purpose!!!

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I actually want to screen what proteins are present in my conditioned medium. But instead of doing a dot blot with the conditioned medium and using different antibodies to test it, I thought it might save a lot more time if I can spot different antibodies onto a membrane and test for all different proteins at the same time, a reversed dot blot if you like. If anyone's got a protocol for doing this I would really appreciate it.

Muly

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The important thing is not how much antibody you can spot on the surface. I want to know which methods you are using to detect Ab-Ag reaction, (fluorescent, QCM, SPR or ....) because depend on the methods you detect, there are different ways to immobilize Ab on the surface.

The other one, which substrate you are using: polymer, Au, Silicon, glass.... each of them also have difference way to modified the surface.

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I'm thinking of using chemiluminescence to detect the Ab-Ag interaction and nitrocellulose membrane as support for the antibodies. I'm only planning to look at a dozen antibodies or so, so will probably be doing manual spotting and want a very simple system to work with.

I'm very new to microarrays and would appreicate any pointer if I'm not going about this in the right way.

Thanks

Muly

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