A new hydrophilic strong/strong type zwitterionic stationary phase for high performance liquid chromatography (HPLC) was synthesized by chemical modification of 3.0 mu m non-porous monodisperse poly(glycidylmethacrylate-co-ethylenedimethacrylate)(P-GMA/EDMA) beads in the following steps. First, the beads were reacted with hydrochloride to obtain chlorizated beads; second, chlorizated beads were reacted with dimethylamine to obtain ammoniated beads; third, ammoniated beads were reacted with 1,3-propanesultone to obtain non-porous hydrophilic zwitterionic stationary phase. The stationary phase was evaluated in detail to determine its ion-exchange properties, separability, reproducibility, hydrophilicity, and the effect of column loading and pH on the separation and retention of proteins. The highest dynamic protein loading capacity of the synthesized zwitterionic packing for bovin serum albumin and Lys were 1.8.3 and 27.4 mg g(-1), respectively. The zwitterionic stationary phase was capable of separating two acidic and three basic proteins simultaneously in less than 2.5 min by the flow-rates of 3.0 mL min(-1). The zwitterionic resin was also used for rapid separation and purification of recombinant human interferon-r (rhIFN-r) and human granulocyte colony-stimulation factor (hG-CSF) from the crude extract solution. The satisfactory results were obtained. (C) 2009 Wiley Periodicals, Inc. J Appl Polym Sci 113: 984-991, 2009