Porous poly(glycidyl methacrylate-co-ethylene dimethacrylate) monoliths with different porous properties grafted with poly(2-acrylamido-2-methyl-1-propanesulfonic acid) chains using cerium(IV) initiated free-radical polymerization have been prepared and used for the separation of proteins in ion-exchange HPLC mode. Because of the presence of the large pores that are typical of monolithic separation media which allow easy flow of all of the mobile phase, the efficiency of the columns does not deteriorate even at high flow velocities as a result of the specific morphology of the monoliths. Optimization of the chromatographic conditions such as the shape of the mobile phase gradient and the flow rate allows for very fast separation of three proteins in less than 1.5 min.