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2 antibodies crossreacting with another protein


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#1 Final year phd at wits end

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Posted 07 May 2010 - 03:58 AM

Hi,

Can ANYONE help me!

I've generated 2 affinity purified antibodies against my protein of interest and BOTH cross-react with another protein.

The antigenic peptides used to generate these antibodies are seperate.
They were injected into the same rabbits but independently affinity purified.
There is no similarity in either antigenic peptide sequence with this other protein.
I cannot think of a reason why two independently affinity purified antibodies would cross-react with the same protein. Does anyone have any suggestions? As I can't explain it!

Thanks

#2 HomeBrew

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Posted 07 May 2010 - 05:12 AM

Could the "other protein" be a different form of the same protein? It's either that, or the "other protein" forms some three-dimensional epitope common to both proteins, but not produced by the same amino acid sequence, or the sequence similarity between the two proteins that form this epitope is not obvious.

If this is polyclonal sera, perhaps you could adsorb out the cross-reacting population?

#3 sgt4boston

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Posted 10 May 2010 - 07:44 AM

I agree with homebrew. Although both your affinity purified abs cross react with another (maybe same) protein what is the level of cross reaction and would you expect this other protein to be in samples you analyze? It could be that the 'cross reactant' is needed at a much higher concentration to produce vaules significant in your test or the cross reactant is not normally present in samples that would be analyzed.




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