New enzymatic assays based on the combination of signal accumulation type of ion sensitive field effect transistor (SA-ISFET) with horseradish peroxidase

Abstract

Peroxidase is widely used for the detection of secondary reactions during measurements of various enzymatic reactions, such as that of oxidase activity, or as an enzyme for immunoassay. Conventional methods utilizing the enzyme require expensive equipment such as a spectrophotometer to measure the absorption of light by the reaction product. Here, we describe a simple and cost-effective method for measuring enzymatic reactions using a signal accumulation type of ion sensitive field effect transistor (SA-ISFET) sensor capable of detecting the proton changes due to the enzymatic reaction. Using this detection principle, we constructed a detection system combining ABTS, an electron mediator, and a horseradish peroxidase activity detection system. As a result, we could quantitatively measure hydrogen peroxide with excellent reproducibility and linearity. As an application of this tool, we describe an oxidase-peroxidase reaction system for the measurement of glucose, sarcosine, uric acid and lactic acid. In addition, we describe an immunoassay system using a peroxidase-labeled antibody for detection of Escherichia coli. We also describe a prototype for a flow-type ISFET device for continuous and routine measurements.