Studies On Novel Amperometric Immunosensors Based On Bilayer Gold Nanoparticles Modified Electrodes

Electrochemical immunosensor has the advantages of immunity technology and electrochemical technology. The detection of the immunoassay is based on the change in the amperometric or potentiometric response before and after antigen-antibody reaction. The shift of response exhibits a linear dependence on the concentration of analytes. This method is highly sensitive, inexpensive, easy-to-use and portable. In this article, we have constructed several immunoelectrodes immobilized antibody onto substrate electrode based on nano-Au and nano-Au/chitosan composite as matrixes. The detailed materials are shown as follows:1. A potentiometric immunosensor for the detection of hepatitis B surface antigen (HBsAg) has been developed by means of electrodeposition and self-assembly techniques. L-cysteine was combined with bilayer gold nanoparticle by its -SH and -NH₂ group. A large amount of hepatitis B surface antibody (HBsAb) was adsorbed by gold nanoparticle and embeded in the PVB firmly because of cage effect of polyvinyl butyral (PVB). Then a potentiometric immunosensor with good sensitivity and stability was prepared. The characteristics of the modified electrode at different stages of modification were studied by cyclic voltammetry (CV). The resulting immunosensor exhibits wide linear range from 8.5 to 256.0 ng.mL^-1 with a detection limit of 3.1 ng'mL^-1. The immunosensor was used to analyze the serum samples with satisfactory results.2. A novel amperometric immunosensor for the detection of hepatitis B surface antigen (HBsAg) has been developed by means of electrodeposition and self-assembly techniques. In this study, L-cysteine was combined with bilayer gold nanoparticle by its -SH and -NH₃⁺. A large amount of hepatitis B surface antibody (HBsAb) was adsorbed by gold nanoparticles. Then an amperometric immunosensor with good sensitivity and stability was made. The characteristics of the modified electrode at different stages of modification was studied by using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), The performances of the immunosensor were studied in detail. The immunosensor showed a specific response to HBsAg in the range 1.28 to 128.0 ng.mL^-1 with a detection limit of 0.32 ng.mL^-1. The correlation coefficient is 0.9982. The immunosensor was used to analyze the serum samples with satisfactory results.3. A novel and sensitive immunosensor has been developed by electro-depositing gold nanoparticles on to a Prussian Blue-modified glassy carbon electrode for determination of hepatitis B surface antigen (HBsAg). After the developed immunosensor was incubated with different concentrations of HBsAg samples at 37℃for 15 min, the current response decreased with an increasing HBsAg concentration in the sample solution. The decreased percentage of the current was proportional to HBsAg concentration ranging from 2 to 300 ng·ml^-1 HBsAg with a detection limit of 0.42 ng·ml^-1 HBsAg (S/N=3). Analytical results of 50 specimens using the developed immunosensor showed satisfactory agreement with those using ELISA, indicating the method to be a promising alternative for detecting HBsAg in clinical diagnosis.4. A sensitive amperometric immunosensor for carcinoembryonic antigen (CEA) was prepared. Firstly, a porous nano-structure gold (NG) film was formed on glassy carbon electrode (GCE) by electrochemical reduction of HAuCl₄ solution, then nano-Au/Chit composite was immobilized onto the electrode because of its excellent membrane-forming ability, and finally the anti-CEA was adsorbed onto the surface of the bilayer gold nanoparticles to construct a anti-CEA/nano-Au/Chit/NG/GCE immunosensnor. The characteristics of the modified electrode at different stages of modification were studied by cyclic voltammetry (CV). In addition, the performances of the immunosensor were studied in detail. The resulting immunosensor offers a high-sensitivity (1310 nA/ng/mL) for the detection of CEA and has good correlation for detection of CEA in the range of 0.2 to 120.0 ng-mL^-1 with a detection limit of 0.06 ng·mL^-1 estimated at a signal-to-noise ratio of 3. The proposed method can detect the CEA through one-step immunoassay and would be valuable for clinical immunoassay.