ENZYME ELECTRODE FOR AMPLIFICATION OF NAD(+) NADH USING GLYCEROL DEHYDROGENASE AND DIAPHORASE WITH AMPEROMETRIC DETECTION/

An enzyme electrode is made from a glassy carbon electrode covered wit h a gelatin membrane containing entrapped glycerol dehydrogenase (GDH) and diaphorase, and protected with a dialysis membrane. Based on ampl ification by the recycling reaction catalyzed by the two-enzyme system s, NAD(+) and NADH can be determined with 800-1200 times higher sensit ivity than for the same electrode in a substrate sensing mode when the flow rate was 0.08 ml/min. The detection limit was about 0.03 mu M fo r NADH. The amplification factors were around 1000 for 0.08 ml/min, wi th quite large variations between electrodes. They had decreased to ab out 70% of the original value after 7 days. The biosensor is intended for detection in immunoassays with alkaline phosphatase as a marker.