Application Of Electrochemistry Immunosensors In DNA Methylation And HIV-1Detection

Electrochemical sensor is an analysis device which can identify, convert signal andfinally realizes detection of bioactive molecules (DNA base small molecules, protein, etc).Combined with chemistry, biology and physics disciplines, it has widely been applied in lifescience, medicine, food and other industries. Graphene, a new carbon material with manyexcellent properties, has wide application prospect in the field of electrochemical sensors. Inthis paper, graphene and self assembly monolayer layer modified electrodes were fabricatedto explore their application in electrochemical sensor. The application of graphene filmmodified electrode in the evalution of DNA methylaiton and self-assembly monolayermodified electrode in protein biosensor were invstigated, enabling simulanteousdetermination of cytosine and5-methylcytosine and the study on the interfacical interaction ofHIV glycoprotein gp120and small molecule inhibitor NBD-556. The main points of thisdissertation are summarized as follows:1. A novel electrochemical sensor for simultaneous determination of5-methylcytosineand cytosine was developed using electrochemically reduced graphene modified glassycarbon electrode. The experimental conditions such as accumulation time, accumulationpotential, pH, were optimized. Under the optimum conditions, the linear range for thesimultaneous determination of cytosine and5-methylcytosine were8~250μmol/L,6~200μmol/L and the low detection limit were0.9,0.7μmol/L, respectively. The electrochemicalsensor exhibited good stability, reproducibility and anti-interference ability. Additionally, theproposed method was successfully applied to the analysis of methylation status in short CpGoligonucleotides with satisfactory results.2. An electrochemical impedance biosensor for investigation the binding interaction ofHIV-1envelope glycoprotein gp120with its small molecule inhibitor NBD-556has beendeveloped. Gp120were immobilized on11-mercaptoundecanoic acid self-assembly monolayersmodified gold electrodes through Au-S interaction. The binding of a specific gp120Phe43cavityto NBD-556recognition layer could be detected by the measurements of the impedance change.The decreased value had a linear relationship with the gp120concentration with a low detectionlimit of36.7ng/mL. The association constants of gp120and NBD-556were calculated to be 0.005(μg/mL)-1(binding affinity1.66μM), comparable to some bio-analysis of3.7μM bindingaffinity, which also demonstrates the feasibility of the method for study the association constantbetween HIV-1glycoprotein gp120and NBD-556.The proposed assay method was rapid, easilyoperated, and also confirmed gp120specific binding capabilities towards the NBD-556on a solidsurface through electrochemical impedance techqiue.