Protein bound to a small piece of DNA will alter the electrophoretic mobility of that DNA fragment. This allows the analysis of protein-DNA interactions, including the measurement of binding rates, affinity, and specificity. In addition, bound and unbound DNA may be isolated from the gel and used for further types of analysis such as methylation interference or uracil interference.

In the mobility-shift (or gel-shift) assay, end labeled DNA is allowed to bind protein. The resulting DNA protein complexes are then run on a non-denaturing PAGE gel and the gel is dried and autoradiographed. The buffer composition of the PAGE gel is varied from the standard TBE gel, because lower ionic strength is needed to facilitate the DNA protein binding. It is interesting to note that binding interactions which can dissociate in free solution within 1 minute show altered mobility on gels which require more than an hour to run. It is theorized that the gel matrix forms a "cage" around the DNA-protein complex, which prevents the components from diffusing away from each other, thus promoting re-association and in effect stabilizing the complex. This would argue that the ratio of bound and unbound material seen on the gel is a direct measure of the fraction of bound material in the sample as it entered the gel. Thus, the conditions set in the binding reaction are of critical importance.

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