Development And Application Of Electrochemical Biosensor Based On Modification Of Au Micro Electrode With Biomolecules

Electrochemical biosensors are devices that combine biosensing interface with electrochemical transducer. With the advantages of high sensitivity, fast analysis, good specificity, simple operation and low cost, electrochemical biosensors have been widely used in clinical diagnosis, food and pharmaceutical analysis, environment monitoring and many other fields. There are several kinds of electrochemical biosensors, each of which differs in fabrication method and detection technology. Enzyme-free electrochemical biosensors and cell biosensors are usually produced by layer-by-layer self-assembly technique, which is time-consuming and lack long-term stability. In this work biomolecules such as chitosan, L-cysteine and folic acid are used to construct enzyme-free biosensors and cell biosensors for environmental monitoring and cancer cell detection.In chapter 1, the development history of electrochemical biosensors, classification of electrochemical biosensors and detection techniques of electrochemical biosensors are reviewed.In chapter 2, an enzyme free electrochemical biosensor based on chitosan and nano gold composite film was developed and applied in nitrite analysis. A layer of gold nanoparticles was first electrodeposited onto gold film micro electrode, followed by the deposition of nano gold and chitosan composite layer, to obtain the GNP@CS/GNP/Au enzyme free electrochemical biosensor. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were used to characterize the electrochemical properties of the modified electrode. It was demonstrated that the electrodeposited gold nanoparticles significantly improved the response signal toward nitrite and the GNP@CS composite membrane accelerated electron transfer and improved affinity toward nitrite ions as well. The detection range of the developed GNP@CS/GNP/Au sensor for nitrite ions was 7.94 μmol/L-4.74 mmol/L, with a detection limit of 4.38 μmol/L (S/N=3). The GNP@CS/GNP/Au electrode was used to determinate nitrite ion in pond water with recoveries ranging from 91.6%-104%, which demonstrated its potential in environmental monitoring.In chapter 3, a cell electrochemical biosensor based on L-cysteine and folic acid (FA) was developed and applied in the determination of HeLa cells. L-cysteine was immobilized onto the surface of gold micro electrode via the interaction of thiol and gold. FA was then bound to the L-Cys/Au electrode with the catalysis of EDC/NHS. BSA was used to reduce nonspecific adsorption on the electrode surface. Based on the strong affinity between FA and FR (Folate Receptor) which is over expressed on HeLa cell membrane, the BSA/FA/L-Cys/Au electrode was used for HeLa cell sensing. The results of EIS and CV characterization demonstrated that the positive charge of L-Cys enhanced the redox reaction on the electrode surface while FA hindered the electron transfer process because of its steric effect. After Hela cells were captured by the FA/L-Cys/Au electrode, the film resistance increased further. The detection range of this constructed FA/L-Cys/Au electrode towards HeLa cells was 1.08x103～1.8×107 cells/mL, with a detection limit of 2.4×102 cells/mL (S/N=3).The FA/L-Cys/Au electrode shows a good reproducibility and good anti-interference performance.