We have the following ELISA situation of serum reacting with antigen
coated plates, washed and followed with biotinylated anti-immunoglobulin.
Then, wash and follow with avidin-HRP. We have one control with serum
followed by avidin-HRP without biotinylated conjugate. We get a strong
signal with the avidin-hrp control. It is clear that avidin-HRP is
reacting with a serum protein bound to the antigen coated plates. We
need antigen coated plates.
The question is: What protein(s) in serum are bound to biotin endogenously?
Is the avidin binding in fact due to a biotin-conjugate? Can avidin
bind via some other ligand interaction independent of biotin, such as
sugars (i.e. lectin-like). If anyone has any answers, please let me
know your ideas.
Basically, I need to find a way to rid of the high non-specific binding
I appreciate aln e-mail response with references, if any. Thanks - Bill