Hemoglobin (Hb) was successfully immobilized on a gold electrode modified with gold nanoparticles (AuNPs) via a molecule bridge 1,6-hexanedithiol (HDT). The AFM images suggested that the HDT/gold electrode could adsorb more AuNPs. UV-vis spectra indicated that Hb on AuNPs/HDT film retained its near-native secondary structures. The electrochemical behaviors of the sensor were characterized with cyclic voltammetric techniques. The resultant electrode displayed an excellent electrocatalytical response to the reduction of hydrogen peroxide (H2O2). The linear relationship existed between the catalytic current and the H2O2 concentration ranging from 5.0 × 10−8 to 1.0 × 10−6 mol · L−1. The detection limit (S/N = 3) was 1.0 × 10−8 mol · L−1.