Nucleic Acid Dye Enhanced Electrochemical Biosensor For The Detection Of Metal Ions

With the development of human industrialization,environment pollution has become more and more serious,which seriously threatens the ecological environment.Heavy metal is a kind of typical pollutant,which poses a serious threat to human health.People pay more attention to their own health condition,therefore,it is important and significant to develop a rapid,simple and reliable detection method for the detection of metal.Electrochemistry is simple,fast,and highly sensitive detection method,which is widely used in the fields of life analysis,environmental analysis and food analysis.In recent years,with the rapid development of biochemistry and nanoscience,different kinds of electrochemical sensors based on biomolecules and nanomaterials have been developed with highly improved selectivity and sensitivity,thus expanding their application range.Two electrochemical biosensors for Hg²⁺and Na⁺were developed combining with nucleic acid dyes as signal enhancers to enhance the current signal on the electrode surface.The main contents of this thesis are as follows.1.A nucleic acid dye-enhanced disposable electrochemical biosensor for the label-free detection of mercury ions(Hg²⁺)was established.The thiolated DNA probes were immobilized to the Au nanoparticles?Au NPs?modified screen printing carbon electrode?SPCE?via Au-s reaction,then Hg²⁺was bound to the DNA probe by thymine?T?-Hg²⁺-thymine?T?mismatch,while Gel Red could be embedded in the double-strand DNA hairpin structure.The current of[Fe?CN?₆]^3-/4-increased due to the reduced electrostatic repulsion between electrode surface and[Fe?CN?₆]^3-/4-;Without Hg²⁺,the interaction between Gel Red and single-stranded DNA is weak,so the current signal of[Fe?CN?₆]^3-/4-kept almost stable.Such a simple strategy provided an sensitive detection of Hg²⁺.Several factors affecting the performance of the sensor were investigated,including electrodeposition time of Au NPs,probe concentration of DNA,incubation time of Hg²⁺,dyes type,concentration of nucleic acid dye,and incubation time of nucleic acid dye.Under the optimal experimental conditions,a good linear relation for Hg²⁺was achieved from 0.1-500 n M with the limit of detection?LOD?of 0.04 n M.Compared with other electrochemical biosensors based on T-Hg²⁺-T mismatch,the sensor is easy to operate and label-free without an amplification procedure.In addition,the developed electrochemical sensor was used to detect two water samples with recoveries ranging from 81%to 114%.The electrochemical biosensor has high sensitivity,and is expected to be used for portable detection of Hg²⁺in water.2.An electrochemical sodium ions?Na⁺?sensor based on Na⁺ DNAzyme and nucleic dye was developed.After modified on the gold electrode surface by Au-S reaction,Ce13d,a DNAzyme,can specifically recognize and bind Na⁺,Ce13d can recognize in the solution,which caused local base folding of Ce13d and thus led to the embedding of SYBR Green I?SG?dye molecules into DNA.As a result,the electrostatic repulsion of the electrode surface of[Fe?CN?₆]^3-/4-reduced and resulted in the current signal increase of[Fe?CN?₆]^3-/4-.Such a strategy provided a simple determination of Na⁺.Furthermore,the structure of Ce13d was studied by fluorescence spectroscopy,and it was found that the changing the number of bases of the stems of Ce13d could affect the confirmation of Ce13d after binding with Na⁺.It is probably because that different base numbers will affect the stability of DNAzyme structure.Electrochemical detection of Na⁺by DPV showed that there was a good linear relationship between peak current of[Fe?CN?₆]^3-/4-and the concentration of Na⁺was linear from 10-50 m M with the LOD was 4.83 m M.The electrochemical Na⁺sensor based on DNAzyme with the advantages of high sensitivity,simple operation,low cost,and label-free is expected to be an appealing method for detection of Na⁺in human blood samples.