The role of reactive oxidizing species (ROS) is proved within numerous physiological processes, including aging, signal transduction and some kind of immune functions. Nowadays ROS and oxidative stress gain increasing attention in connection with a wide spectrum of diseases. In case of Asian people, the enzyme taking part in the ethanol metabolism, the aldehyde dehydrogenase is absent or mutated, that can result in liver tissue damage upon extensive alcohol consumption. Most of the ROS species are electrochemically active therefore the applications of electrochemical methods are the most promising for in situ or in vivo monitoring or quantification of them. In our work development and improvements of selective and sensitive method for electrochemical detection of these molecules and radicals are attempted. We prepared ultra thin size-exclusion layer by electropolymerization of m-phenylenediamine monomer on the surface of the Pt working electrode to ensure its selectivity. We have worked out the optimal circumstances for the selective layer preparation and tested its stability and function. In order to enhance the sensitivity of ROS detection a new amperometric method, the periodically interrupted amperometry (PIA) was developed and applied. With this approach we succeeded selective and sensitive detection of H2O2 in vitro.