Hi,

has anyone experience with EMSA ? how to say that the shift is specific ? I tried with competitors , but the band is disapearing but not completely with 100x of competitor (should it).... and it is also slowly disapearing with addition of mor pdIdC.
And there is also a problem with supershift, the first time I try with ab, I got a supershift but now I'm trying to repeat it, but there is no supershift anymore, just a shift ???

The best way to show specificity is with a supershift. Competition and mutant oligos can complement this work. Supershifts may work under different conditions, such as preincubation with antibody, or post oligo binding incubation with antibody may work best. Sometimes incubation on ice is also necessary.
IF your band is disappearing with poly-didc this is a serious problem. Your bands are certainly non-specific. All oligos can bind some protein or another. Therefore mutating specific sites aand supershifting is important.