This paper describes an electrochemical immunosensor for simple, fast and quantitative detection of a urinary hippuric acid which is one of major biological indicator in toluene-exposed humans. The feature of this electrochemical system for immunoassay of hippuric acid is based on the direct conjugation of ferrocene to a hippuric acid. With the competition between the ferrocene-hippuric acid complex and hippuric acid for binding to the anti-hippuric acid monoclonal antibody coated onto gold nanoparticles, the electrical signals are turned out to be proportional to urinary hippuric acid in the range of 0.01-10 mg/mL, which is enough to be used for the point-of-care. The proposed electrochemical method could extend its applications to detect a wide range of different small molecules of antigens in the health care area.