HIS elution: can imidazole be added after SDS-sample buffer? - (Aug/06/2012 )
I was doing some pull-down and IP and it seems I treated all samples equally and instead of doing an elution step I directly added SDS-sample buffer containing B-mercaptoethanol to "realease" the complexes from the beads.
The problem is that some of the pull downs were HIS-tag proteins (I used HIS-tag dynabeads from invitrogen) but I did not add any imidazol. When I have realised of this, I had already added the SDS sample buffer and heated the samples (95C, 5min). It is possible to add the imidazole after this or SDS+B-mercapto+boiling it enough to release the complexes?
Thanks a lot,
sds-bme-boiling should release the complexes and denature them. adding imidazole will make no difference at this stage.