A novel strategy to construct a sensitive amperometric sensor of H2O2 was described. Horseradish peroxidase (HRP) wassuccessfully immobilized on nanometer-sized Au colloids, which were supported by thiol-tailed groups of cysteaminemonolayer. The thiol-tailed groups were formed through the covalent binding of glutaraldehyde on a cysteamine-modifedgold electrode. With the aid of the catechol mediator in the solution, HRP-labeled Au colloids displayed excellentelectrocatalytical response to the reduction of H2O2, which increased with decreasing size of Au colloids. The sensorsresponded to H2O2 in the concentration range of 0.39 mM-0.33 mM, and reached 95% of the steady-state current in less than5 s with the detection limit of 0.15 mM. The response showed a Michaelis-Menten behavior at larger H2O2 concentrations.The KappM values for the sensors based on different sized HRP-labeled Au colloids (colloid A-D) were found to be 0.11, 0.094,0.054 and 0.064 mM, respectively. The low KappM values demonstrated that HRP immobilized on Au colloids exhibited a highaf