Sandwich ELISA from direct? - (May/18/2011 )
Experience: somewhat familiar with ELISAs
Question: Can I create a sandwich ELISA from a direct ELISA method?
Background: I am trying to detect/quantify a unique transgenic protein that i'm extracting from plant tissue. The problem is the only method for immuno-detection is a direct ELISA (for pure bacterial produced) which requires I bind the protein (antigen) to the plate and then add a MAb specific for the antigen, followed by a secondary antibody for the MAb.
Problem: I cannot get a signal when the plant-soluble protein is bound as a whole to the plate. I would like to develop a sandwich ELISA from the materials I have if possible so that I can add a 'dirty sample' and get good detection. Is this possible???
Thanks thanks thanks
you will need a second antibody to your antigen. it should be from a species not recognized by your secondary antibody and should be against a different epitope than your antibody. coat the plate with this antibody.