Abstract: A dispersion of multi-wall carbon nanotubes (MWCNT′s) in polydipropylenedimethylammonium chloride (PDDA) was added dropwise onto the surface of glassy carbon electrode (GCE) to prepare modified electrode of PDDA-MWCNT′s/GCE (abbr. as electrode Ⅰ); gold nanoparticles (AuNP′s) were plated on the surface of electrode Ⅰ by constant potential electro-deposition, the modified electrode of AuNP′s/PDDA-MWCNT′s/GCE (abbr. as electrode Ⅱ) was obtained. Haemoglobin (Hb) was dropped and immobilized on the surface of electrode Ⅱ to obtain the biosensor Hb/AuNP′s/PDDA-MWCNT′s/GCE (abbr. as electrode Ⅲ) which after rinsing with PBS of pH 6.8, was used for determination of H2O2. As compared with bare GCE, electrode Ⅰ and electrode Ⅱ, in a PBS of pH 6.8, the reduction peak current of H2O2 at electrode Ⅲ was significantly increased. Linear relationship was kept between values of reduction peak current and concentration of H2O2 in the range of 1.0 to 1 800 μmol·L-1, with detection limit (3S/N) of 0.8 μmol·L-1. In the application of this biosensor to the determination of H2O2 in a disinfectant, results obtained by this method were in consistency with those found by the KMnO4 titrimetric method.