An electrochemical platform for parallel monitoring of secreted alkaline phosphatase (SEAP) has been microfabricated on a device with a mammalian-cell array chip. A 4 × 4 ring–ring electrode array was designed at the rim of the round cellular pattern with a diameter of 270 μm. Electrochemical characterization was carried out, and it was found that the collection efficiency was about 50% in dual mode when the inner-ring and the outer-ring electrodes were selected as the collector and generator electrodes, respectively. The current amplification ratio for the dual mode normal to single mode was 2.84. SEAP expressing from the cells was parallelly monitored by using a multiplexer switching system at the 16 round cellular spots. The reduction current for HeLa cells transfected with plasmid encoding SEAP observed at the collector outer ring electrode was found to be significantly higher than that for wild-type HeLa. Finally, the top of the microwell with the round cellular pattern was covered with a poly(dimethylsiloxane) block for 5 min to accumulate the secreted enzyme and the product of the enzyme reaction so that further signal enhancement could be observed.