Hi to all, I'd like to perform an immunoblotting against a protein on my EMSA. I would like to know if I can strip the streptavidin-HRP / biotinylated DNA probe complex from the membrane and re-probe it with the specific antibody. Thanks .
It may be easier to use a different chemistry for your secondary antibody, such as alkaline phosphatase or if you have access to a LiCor system.
Otherwise you would be looking at stripping the whole DNA probe complex from the membrane (Strep-HRP will never dissociate from biotin probe). I presume this is a nylon membrane, so you could try stripping buffers designed for Northern/ Southern (try Thermo Scientific website for the protocols), but I wouldn't be too confident that your protein would remain on the membrane after these harsh treatments.
You could try to kill the remaining HRP activity with sodium azide or excess of H2O2 then wash an reprobe with your antibody.
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