One-step separation-free amperometric biosensor for the detection of protein

A novel enzyme biosensor for the detection of protein is presented. The bio sensor was made from a screen-printed three-electrode configuration. Amino acid oxidase was immobilized with glutaraldehyde and polyethylenimine on a working electrode made of rodinised carbon. A protease was immobilized on a n immunodyne membrane and was placed on the electrode. A protein sample was deposited on the membrane, and was subsequently hydrolyzed to amino acids in the presence of the protease. This in turn produced hydrogen peroxide by the immobilized amino acid oxidase. The oxidation of hydrogen peroxide was then detected at +400 mV vs. an Ag/AgCl reference electrode. The method wa s very effective at detecting a very low level of protein. The sensor does not require any washing step. The sensor works with only 40 mu l of sample per detection, and may be used on-site as a disposable sensor using a hand- held meter. The electrodes are also stable for more than 6 weeks. (C) 2000 Elsevier Science B.V. All rights reserved.