is there any problem if the fusion protein was used directly as the antigen? - fusion protein was used directly as the antigen (Apr/30/2006 )
Hi, dear friends:
I have expressed a protein in the pET-32a system and plan to use it as the antigen to immune rabbits. But because my cloned gene was inserted into the NcoI&SalI site after double digestion, the expressed protein is acutually a fusion of the tag(including the trx tag, the his tag, the s tag etc.) and the foreign gene. So could anyone experienced tell me is it feasible to use the fusion protein directly as the antigen? My purpose is very simple, to produce antibodies for further western blot analysis of my cloned gene.
If you want to make anti-tag antibodies, yes.
You might produce more easily anti-tag antibodies than anti-protein antibodies.
Thank you very much, dear missele!
I have successfully made antibodies to a his-tagged protein to use in westerns without problems. Supposedly the His-tag is not very immunogenic. Im not sure about other tags.
depends on whether the tag is buried when the protein folds