How is the Thermo Scientific Pierce Direct IP Kit method different from traditional immunoprecipitation?
Traditional immunoprecipitation involves incubation of an antibody with a sample that contains the protein antigen of interest, followed by capture of the antibody:antigen complex to immobilized Protein A or Protein G beaded agarose resin. Finally, after washing to remove nonbound (presumably undesired) components of the sample, the antigen and antibody are recovered by boiling the resin in denaturing sample loading buffer for analysis by SDS-PAGE. The traditional procedure is performed in a microcentrifuge tube, requiring that the solution be carefully withdrawn from the agarose resin after they have been pelleted by centrifugation. In the very least, both the target antigen and the antibody are eluted in the final fraction, resulting in the presence of prominent heavy and light chain antibody bands in the final electrophoresis gel. If the target protein has a similar molecular weight to one of the contaminating antibody fragments, its presence may be masked, making it impossible to i
