A microfluidic device that forms a row of liquid plugs in a micro flow channel was fabricated, and its applicability to an on-chip enzyme-linked immunosorbent assay (ELISA) was demonstrated. The nanoliter liquid plugs were formed by six independent pumps and were mobilized in the micro-flow channel using a main pump. The operation of the pumps was based on the volume change caused as a result of the electrochemical production of hydrogen bubbles. When the bubbles were produced in the pumps, a polydimethylsiloxane (PDMS) diaphragm at the bottom of the compartment inflated and closed an inlet hole at the top of the reservoir. The solution in the reservoir was forced to be injected into the flow channel forming a liquid plug. After six plugs were formed, they were mobilized to a reaction chamber one by one to allow an antigen-antibody binding, cleansing, and detection. This simple and reliable liquid handling system could be applied to the detection of a tumor maker, oc-fetoprotein (AFP).