Electrophoretic exclusion, a novel separations technique that differentiates species in bulk solution using the opposing forces of electrophoretic velocity and hydrodynamic flow, has been adapted to a microscale device. Proof-of-principle experiments indicate that the device was able to exclude small particles (1 mu m polystyrene microspheres) and fluorescent dye molecules (rhodamine 123) from the entrance of a channel. Additionally, differentiation of the rhodamine 123 and polystyrene spheres was demonstrated. The current studies focus on the direct observation of the electrophoretic exclusion behavior on a microchip.