can protein modifications affect immunodetection?

[Jayakumar, R]

All three are strong possibilities.  C is probably right. You should try developing monoclonal antibodies with recombinant protein and screening them for specificity against the native form.
  To check "b", try Immunoprecipitating the protein with the antibody.  

Jay
 

-----Original Message-----
From: methods-bounces from oat.bio.indiana.edu [mailto:methods-bounces from oat.bio.indiana.edu] On Behalf Of Scientist
Sent: Wednesday, July 22, 2009 1:25 PM
To: methods from magpie.bio.indiana.edu
Subject: can protein modifications affect immunodetection?

Hi all!
im trying to detect a  protein in plants Crude extract using polyclonal Abs. These Abs were obtained from rat sorum, The protein used to innolculate the rats was expressed in E.coli (thus lacking the typical postrasductional modifications occurring in plants) .
Untill only no positve results were obtained.
When recombinant form is used as positive control everthing works well.
So the possibilities are :
a) native form is absent in the plant.
b) the protien is present but in low concetration
c) native form is modified in someway , thus the abs cannot recognize the epitopes.

have anyone a suggestions for these items?
thanks a lot
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