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Western Blot Antibody Epitope

antibody western blot epitope

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#1 southtowns18

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Posted 06 June 2012 - 07:35 PM

I have a question concerning an antibody I'm using. The antibody was generated by immunizing a mouse with a peptide corresponding to the n-terminal residues 2-20 of the protein I'm looking at. Unfortunately, the lab that generated this Ab does not list the specific sequence.

The N-terminal residues 1-20 of the protein I'm looking at correspond to the signal sequence. Does that mean the antibody I'm using detects the signal sequence of this protein? I thought the signal sequence gets cleaved off.

Or does "residues 2-20" mean residues 2-20 after the signal sequence gets cleaved off (so residues 22-40 if your counting the signal sequence). For example, other Abs I use to detect this protein might say the epitope is residues 40-60 from one company but 60-80 from another company depending on whether they include the signal sequence. But bc these companies list the exact sequence I can tell that they are the same and find exactly where it lies within the protein.

Obviously I wouldnt need to ask this if the lab who made it just listed the exact sequence but whatever. Hopefully someone can help me with this issue. Thanks in advance and hopefully my wording didnt make it too complicated.

#2 bob1

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Posted 08 June 2012 - 01:42 PM

Without knowing which antibody you are using, it is impossible to say. One way you could test it is to run a western - if you get a band corresponding to you protein of interest, running at about the right size, then it will be against the cleaved protein, but if you get a band running at about 2 kDa, then you are detecting the cleaved product.





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