Amperometric enzyme immunosensors based on layer-by-layer assembly of gold nanoparticles and thionine on Nafion modified electrode surface for α-1-fetoprotein determinations

Abstract

A novel amperometric enzyme immunosensor with amplified sensitivity has been developed for the detection of α-1-fetoprotein (AFP) based on layer-by-layer (LBL) assembly of gold nanoparticles (nano-Au) and thionine (Thi) on Nafion (Nf) modified electrode surface by electrostatic adsorption. The stepwise layer-by-layer adsorption interface morphology was further characterized by means of electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The enzyme, horseradish peroxidase (HRP), was employed to block the possible remaining active sites of the nano-Au monolayer, avoid the non-specific adsorption instead of bovine serum albumin (BSA) and amplify the response of the antigen–antibody reaction. The performance and factors influencing the performance of the resulting immunosensors were studied in detail. Tests performed with this immunosensor showed good linearity and specificity when it was evaluated on several standard serum samples and the working range for the system was 1 to 250 ng/ml with a detection limit of 0.56 ng/ml. In addition, the prepared immunosensor could be regenerated 8 times with 4 M urea solution. When the immunosensor was stored at 4 °C and measured intermittently (every 3–5 days), no apparent change was found over 3 months, it yielded a 6.5% R.S.D.