Hemoglobin (Hb) was immobilized in poly-allylamine (PAA) film onto the gold electrode by layer by layer (LBL) method. The modified electrode exhibited a pair of well-defined peaks during cyclic voltammetry, which was attributed from the direct electron transfer of heme proteins. The immobilized Hb showed an excellent electrocatalytical response to the reduction of hydrogen peroxide. The sensor exhibited a fast response and high sensitivity. Through the use of optimized conditions, the linear range for H2O2 detection was from 2.5 x 10(-6) M to 5 x 10(-4) M with detection limit of 0.2 mu M. The proposed biosensor showed long-lasting stability and excellent reproducibility. (c) 2006 Elsevier B.V. All rights reserved.