I have been using kits with paired monoclonal antibodies, what I wish to do is purchase polyclonal antibodies to a particular antigen and split in two, immobilise one half and fluoro label the second half.
Is this a feasible idea?
yes, of course, as long as you don't use labelled antibodies to these polyclonal antibodies for detection.
In principle that should work. It's suggested to use polyclonal ab for capturing at and monoclonal ab for detecting antigen. However it's a matter of concern if ag is detected in defined system or in clinical specimen plus animal species in which poly- & mono- clonal and r raised.
Will depend on generation of antibody against different epitopes present in the antigen. During polyclonal antibody development the epitope which has more foreign nature will produce high affinity and more amount of antibody in the pool as compare to other epitopes. Therefore, using only same polyclonal antibody for sandwich assay purpose is a tricky one. It may work or may not work.
With best wishes.
It's depend of the immunogenicity of your antigen to whom antiserum was elicited. If your antiserum (once purified) become quite diverse against multiples epitopes on the antigen, with similar affinity over all of them, probably your proposed should be work. Otherwise if your antiserum has a poorly epitopes recognition, not properly oriented and/or not repetitive enough on the antigen, you probably will front problems of hindrance, competition between the catcher and the labelled, and finally a poor sensitivity in the response of the assay.
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