The preparation of chemically modified gold electrodes by thionine covalently bound to self-assembled cysteamine monolayer (SAM) is described. This thionine-SAM modified electrode is electroactive and exhibits two new electrochemical redox couples with formal potentials which are more positive than that of thionine in bulk solution. An obvious electrocatalytic oxidation of nicotinamine adenine dinucleotide (NADH) is observed; meanwhile, the anodic overpotential is reduced by about 600 mV compared with the results obtained at bare gold electrodes. The participation numbers of H+ for the electroactive SAM are 2 and 1 corresponding to the two electro-couples I and II respectively. The apparent electron transfer rate constant is about 0.02 s(-1) at pH 5.0-9.0 for the redox-couple 1. The amperometric response to NADH increased linearly in the range of 2.0 X 10(-5) to 1.0 x 10(-2) M. The electrolysis results show that the oxidation of NADH is a two-electron process and the product formed is enzymatic active NAD(+). In addition, this thionine self-assembled modified electrode shows good stability for long-term use.