The formation of horseradish peroxidase-lipid Langmuir-Blodgett film and its applicability as a biosensor have been studied. HRP was spread directly onto the subphase covered with a layer of lipid in LB trough. Our experimental results showed that surface pressure of this film from liquid to solid state ranged between 15 to 25 mN/m. At surface pressure of 25 mN/m, the monolayer was successfully transferred onto the gold surface. In addition, electrochemical properties of this film showed that protein molecules still kept their natural structure and can give a well electrocatalytic activity to H2O2. As an H2O2 biosensor, this LB film was able to detect concentration of H2O2 which were 3 x 10(-7) M. (c) 2007 Elsevier B.V. All rights reserved.