An amperometric enzyme electrode based on peroxide detection has been prepared for protein determination. A trienzyme system immobilized on two stacked membranes was used. L-aminoacid oxidase and leucine aminopeptidase were coimmobilized on one membrane in contact with the platinum anode, whereas the second membrane, directly in contact with the reaction medium, had immobilized trypsin. In a first step, the sensor equipped only with the bienzyme membrane was tested with two peptides, then with protein hydrolysates, and optimized conditions were determined. Casein could be assayed with the three-enzyme system within a linear concentration range of 0.1–4 mg/mL with response-times between 5 and 9 minutes.