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A novel H(2)O(2) biosensor is described which is based on immobilization of horseradish peroxidase (HRP) on DNA/electrodeposited, ZrO(2)/modified, gold electrode. The DNA is attached via its 5' end to ZrO(2) and this provides a microenvironment for the immobilization of various biomolecules and promotes electron transfer between HRP and the electrode surface. Under optimized conditions, the biosensor reduced H(2)O(2) linearly between 3.5 microM and 10 mM with a detection limit of 0.8 microM at a signal-to-noise ratio of 3. In addition, the developed biosensor shows an acceptable stability and repeatability. Importantly, the analytical methodology could be further developed for the immobilization of other proteins and biocompounds.

Alternate adsorption of positively charged colloid-Au nanoparticles (nano-Au ⊕ ) and negatively charged hemoglobin (Hb) on L-cysteine (L-cys) modified gold electrode resulted in the assembly of {Hb/nano-Au ⊕ } n layer-by-layer films/L-cys modified gold electrode. The nano-Au ⊕ was characterized by transmission electron micrograph (TEM) and microelectrophoresis. The modified electrode interface morphology was characterized by electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM), cyclic voltammograms (CV) and chronoamperometry. Direct electron transfer between hemoglobin and gold electrodes was studied, and the apparent Michaelis-Menten constant ( app m k ) of the modified electrode was evaluated to be 0.10 mmol·L −1 . Moreover, the higher activity of proteins in the nano-Au ⊕ films could be retained compared with the electropolymerization membrane, since the proteins in nano-Au ⊕ films retained their near-native structure. Direct electron transfer between hemoglobin and electrode and electrochemically catalyzed reduction of hydrogen peroxide on a modified electrode was studied, and the linear range was from 2.1×10 −8 to 1.2 ×10 −3 mol·L −1 (r = 0.994) with a detection limit of 1.1×10 −8 mol·L −1 H 2 O 2 .positively charged colloid-Au nanoparticles (nano-Au ⊕ ), hemoglobin, catalyze, hydrogen peroxide