In this paper, the hemoglobin (Hb)-collagen microbelt modified electrode with three-dimensional configuration was fabricated via the electrospinning method. Direct electron transfer of the Hb immobilized into the electrospun collagen microbelts was greatly facilitated. The apparent heterogeneous electron transfer rate constant (k(s)) was calculated to be 270.6s(-1). The electrospun Hb-collagen microbelt modified electrode showed an excellent bioelectrocatalytic activity toward the reduction of H(2)O(2). The amperometric response of the biosensor varied linearly with the H(2)O(2) concentration ranging from 5 x 10(-6) mol L(-1) to 30 x 10(-6) mol L(-1), with a detection limit of 0.37 x 10(-6) mol L(-1) (signal-to-noise ratio of 3). The apparent Michaelis-Menten constant (K(m)(app)) was 77.7 mu mol L(-1). The established biosensor exhibited fast amperometric response, high sensitivity, good reproducibility and stability. (C) 2011 Elsevier B.V. All rights reserved.