The sensitivity of many diagnostic immunoassays is often determined by the final concentrations of the analytes detected. In diagnostic applications that involve a binding immununoassay, the final concentration of the detected analyte may be increased by enhancing the binding reaction or concentrating the immuno-complex before detection. In this study, we use isotachophoresis (ITP) to enhance the binding reaction of an immunoassay as well as to concentrate the complex of interest. We demonstrate the above idea in a microchip-based, liquid-phase alpha-fetoprotein(AFP) immunoassay, which can be used as a marker for liver cancer. In this system, we react AFP with a DNA-labeled antibody and a fluorescent dye-labeled antibody producing a sandwich immuno-complex with AFP. We enhance the reaction by pre-concentrating the DNA-labeled antibody using ITP. The complex of interest is also concentrated using ITP to enhance the detection sensitivity. The concentrated complex is separated using Gel Electrophoresis (GE) in an integrated microchip system that automates the pre-concentration, reaction and separation steps. We demonstrated a better than 1pM limit of detection of AFP using experiments and simulations.