Electrochemical immunosensor for norethisterone based on signal amplification strategy of graphene sheets and multienzyme functionalized mesoporous silica nanoparticles

Norethisterone is one kind of widely used anabolic steroid hormones which can help to promote livestock growth and sometime has been illegally used for livestock breeding. The residues of norethisterone in animal food will harm people's health, therefore, it has been banned to use for the growth promotion purposes in livestock. In this study, amino-group functionalized mesoporous silica nanoparticles (MSN) were prepared and used to immobilize Au nanoparticles, which was further utilized for the adsorption of horseradish peroxidase (HRP) and the secondary antibody (Ab(2)). The resulting nanoparticles, Au-MSN-HRP-Ab(2) were used as labels for immunosensors to detect norethisterone antigen. A sandwich-type protocol was used to prepare the immunosensor with the primary antibody (Ab(1)) immobilized onto thionine (TH) and graphene sheet (GS) modified glassy carbon electrode surface. The sensitivity of the sandwich-type immunosensor using Au-MSN-HRP-Ab(2) as labels for norethisterone antigen detection was much higher than that using either Au-MSN-Ab(2) or MSN-HRP-Ab(2) as labels. Within norethisterone concentration range (0.01-10 ng/mL), a linear calibration plot (Y=0.55671 + 8.27101X, r=0.9993) was obtained with a detection limit of 3.58 pg/mL under optimal conditions. The proposed immunosensor shows good reproducibility, selectivity, and acceptable stability. This new type of labels for immunosensors may provide many potential applications for the detection of growth hormone in animal derived food. (C) 2010 Elsevier B.V. All rights reserved.