Binding using Biotinylated Poly Ab and Protein with His-tag

Dear SPR,

I have a request to do KD analysis using a Biotinylated Poly Ab with a Protein with a Histag (labeled on C terminal). Planning on immobilizing the Biotin-poly-Ab using a Biacore strepavidin chip. And pass on the Protein with Histag. In theory, would this work? Can the poly Ab recognize the Protein even with a Histag?. Thanks.

Since you use a polyclonal antibody to capture your His-tagged protein I would not be worried by the His-tag. You can check this by coating a little of you His-tagged protein in an ELISA plate and detect it with the biotinylated poly ab and a suitable secondary antibody with HRP or strept-HRP.

Arnoud

Thanks Arnoud for your reply. A follow up question. I was planning on doing a multicycle kinetics but have no idea how the regeneration will come out using a typical regen solution that I have been using (ie. glycine 1.5). Hoping not to spend time on method development for this Regeneration. Would a single cycle be an option to begin with? I needed to see binding first, if I see it using single cycle, then perhaps I can devote time on the multi-cycle. Kindly comment. Thanks.

Single cycle kinetics also needs a regeneration at the end. When this regeneration fails, following injections will be less accurate.
You can use the sck to quickly optimize the analyte range but not to avoid regeneration optimazation.
Arnoud