Electromobility focusing (EMF) is a relatively new protein separation technique that utilizes an electric field gradient and a hydrodynamic flow. Proteins are focused in order of electrophoretic mobility at points where their electrophoretic migration velocities balance the hydrodynamic flow velocity. Steady state bands are formed along the separation channel when equilibrium is reached. Further separation and detection can be easily achieved by changing the electric field profile. In this paper. we describe an EMF system with on-line UV absorption detection in which the electric field gradient was formed using a dialysis hollow fiber. Protein focusing and preconcentration were performed with this system. Voltage-controlled separation was demonstrated using bovine serum albumin and myoglobin as model proteins. The limitations of the current method are discussed, and possible solutions are proposed.