Abstract
BACKGROUND: The purpose of this study was to devise a novel electrochemical immunosensor for ultrasensitive detection of alfa-fetoprotein based on Fe(3)O(4)/Au nanoparticles as a carrier using a multienzyme amplification strategy. METHODS AND RESULTS: Greatly enhanced sensitivity was achieved using bioconjugates containing horseradish peroxidase (HRP) and a secondary antibody (Ab(2)) linked to Fe(3)O(4)/Au nanoparticles (Fe(3)O(4)/Au-HRP-Ab(2)) at a high HRP/Ab(2) ratio. After a sandwich immunoreaction, the Fe(3)O(4)/Au-HRP-Ab(2) captured on the electrode surface produced an amplified electrocatalytic response by reduction of enzymatically oxidized hydroquinone in the presence of hydrogen peroxide. The high content of HRP in the Fe(3)O(4)/Au-HRP-Ab(2) could greatly amplify the electrochemical signal. Under optimal conditions, the reduction current increased with increasing alfa-fetoprotein concentration in the sample, and exhibited a dynamic range of 0.005-10 ng/mL with a detection limit of 3 pg/mL. CONCLUSION: The amplified immunoassay developed in this work shows good precision, acceptable stability, and reproducibility, and can be used for detection of alfa-fetoprotein in real samples, so provides a potential alternative tool for detection of protein in the laboratory. Furthermore, this immunosensor could be regenerated by simply using an external magnetic field.