Research On The Construction Of Photoelectrochemical Biosensors Based On The Mutiple Signal Amplification Strategies

Photoelectrochemistry（PEC）used excitation light at an appropriate wavelength to excite the substance（generally the photoactive material modified on the photoelectrode）to generate electron transfer,and then directly or indirectly detected the changes of the photocurrent through appropriate methods to realize the detection of analyte.The difference between PEC and chemiluminescence or electrochemistry was that in its analytical system,not only the photoactive materials were required to have the photoelectric effect,which could produce photocurrent signal,but also the analyte was required to participate directly or indirectly in the reaction process,causing the change of photocurrent signal,so as to achieve the purpose of detection.In this process,signal amplification strategy was an indispensable factor.Combining the appropriate signal amplification strategy to amplify the photocurrent signal was conducive to further analysis and detection so as to effectively improve the detection sensitivity of sensor.Therefore,this paper was mainly based on some environmentally friendly and stable photoactive materials,combining some appropriate signal amplification strategies in PEC analysis,so as to improve the analysis performance of the designed biosensors and achieve sensitive detection of the analytes.The main research contents are as follows:1.A PEC biosensor based on the sensitization strategyIn PEC analysis,some organic dyes can be embedded in double-stranded DNA（dsDNA）,and used as the sensitizer of the basic photoactive matrix to form a sensitized structure,achieving the signal amplification.Commonly used sensitizers such as[Ru（dcbpy）₂dppz]²⁺can be stably embedded into dsDNA,but it is not conducive to biological analysis due to the high toxicity and high cost.Therefore,it is necessary to find some sensitizers with low toxicity,low cost and can be stably embedded in dsDNA.Besides,the common short dsDNA structure is often used to realize the immobilization of sensitizer,which greatly limits the amount of immobilization of sensitizer,leading to low sensitization efficiency.Doxorubicin（Dox）can be stably embedded in dsDNA due to its structural characteristics and it is low toxic.Based on this,Dox was supposed to be used as a sensitizer of g-C₃N₄,the photoelectric conversion efficiency of g-C₃N₄ was improved due to the formation of a sensitization structure.At the same time,the target（micro RNA-141）activated DNA walker propelled by catalytic hairpin assembly（CHA）could trigger the hybridization chain reaction（HCR）to produce numerous long dsDNA copolymer complexes,which effectively increased amount of immobilization of Dox and greatly increased its sensitization efficiency.The proposed PEC biosensor exhibited a linear range from 0.25 fmol·L^-1 to 2.5 nmol·L^-1 with a detection limit of 83 amol·L^-1 and offered a simple and effective approach to detect biomarkers.2.A PEC biosensor based on the steric hindrance effectIn the previous work,although the signal amplification was realized by the combination of organic dye sensitizer and inorganic semiconductor material to form a sensitized structure,it was often necessary to provide a large immobilization platform for the sensitizer through the complex electrode preparation process,which could result in more experimental steps and longer analysis time.Therefore,it is of great significance to build a simple and effective PEC biosensor with the help of appropriate signal amplification strategy.Herein,a PEC biosensor for target（a DNA fragment of HPV-16）detection was successfully constructed based on iron oxyhydroxide（Fe OOH）which was used as the photoactive material and exonuclease III（Exo III）-aided dual recycling signal amplification strategy.Firstly,Fe OOH could firstly generate a high PEC signal.Then the Exo III-aided dual recycling signal amplification strategy could convert a trace amount of target into numerous single-stranded DNA（S1）and further introduce substantial signal probe DNA labeled-Si O₂ NPs（SP-Si O₂ NPs）on the electrode surface,thus the PEC signal could significantly decrease owing to the steric hindrance effect of Si O₂ NPs.The constructed PEC biosensor exhibited a good analytical performance from 0.5 fmol·L^-1 to1 nmol·L^-1 with a detection limit of 0.17 fmol·L^-1.Importantly,this work provided a reliable tool for the detection of various biomarkers and exhibited a potential application in bioanalysis and disease diagnosis.3.A PEC biosensor based on the p-n semiconductor quenching effect and enzymatic catalytic precipitation reactionIn the previous works,although the PEC biosensors were successfully constructed with the help of appropriate signal amplification strategies,they belonged to a single signal amplification strategy in essence.Therefore,it is envisaged whether the analytical performance of the biosensor can be further improved by combining two or more signal amplification strategies by using the synergistic effect of each other.In this experiment,we found that copper bismuth（Cu Bi₂O₄,CBO）possessed the intrinsic peroxidase-like activity.Besides,it is a p-type semiconductor.Herein,n-type semiconductor 3,4,9,10-perylenetetracarboxylic dianhydride（PTCDA）was used as a basic photoactive matrix.At the same time,with the help of Nt.A1w I enzyme-assisted target（a DNA fragment of p53gene）recycling amplification and catalytic hairpin self-assembly,CBO could be introduced to the electrode surface through the specific binding of biotin and streptavidin.Because CBO could cause the p-n semiconductor quenching effect and enzymatic catalytic precipitation reaction,the photocurrent signal could be quenched subsequently.The biosensor had a good response signal with the concentration of target range from 1fmol·L^-1 to 1 nmol·L^-1 and a detection limit of 0.33 fmol·L^-1.Besides,this biosensor provided a new idea for exploring other multifunctional semiconductors with simulated enzyme activity.