Photopolymerization-Based Electrochemical Biosensor And Its Ultrasensitive Detection Of Cancer-Related Nucleic Acids

Cancer is one of the main causes that seriously threatens mankind health and even leads to death,and early detection and treatment is the most effective means to improve the survival rate of cancer patients,thus the realization of early cancer screening is an urgent problem to be solved.Biomarker of cancer nucleic acids can provide an important basis in vitro diagnostic for early cancer screening;it is of great importance to develop a strategy of trace nucleic acids detection with high specificity,simple and rapid,and high sensitivity for in vitro diagnosis of cancer due to the small sequence difference,poor stability and low expression of nucleic acids.Electrochemical analysis technique has the advantages of high sensitivity,fast detection and low cost,so it is expected to be developed into a portable nucleic acid analysis platform.However,the existing electrochemical detection methods still have some defects,such as long detection cycle,complex construction process,insufficient sensitivity and high cost.The intrinsic chain dynamic growth process of free radical polymerization can link the monomer labeled by electroactive probes into macromolecular chains,and directly transform the recognition event of a single nucleic acid into the electrochemical response of numerous electroactive probes,which can realize the detection signal amplification.Thus,this paper establishes a novel method based on electrochemical signal amplification with the help of photoinduced free radical polymerization,and constructs a variety of electrochemical biosensors around different kinds of photopolymerization for the ultrasensitive detection of cancer-related nucleic acids.The research contents are as follows:（1）Cu Fe₂O₄-Enhanced Photopolymerization Amplification Strategy for Quantitative Determination of Lung Cancer-Related Nucleic AcidsAn electrochemical biosensor based on Cu Fe₂O₄ enhanced photoinitiated atom transfer radical polymerization（ATRP）is constructed for ultrasensitive detection of lung cancer-related nucleic acids in this research.Peptide nucleic acids（PNA）is chosen as the capture probe for target DNA,and ATRP initiator is introduced to the electrode surface via the coordination of phosphate-Zr⁴⁺-carboxylate.The reduction quenching cycle mediated by copper（II）complex and the Fenton-like reaction of Cu Fe₂O₄-H₂O₂ are simultaneously triggered under the irradiation to initiate free radical polymerization.Therefore,the polymeric chains begin to grow under polymerization regulated by those two systems,and a large amount of electroactive monomer are grafted from the electrode surface.The limit of detection is as low as 1.98 a M（R²=0.998）and the detection range is from 0.1 f M to 10 p M.（2）Cu₂（OH）PO₄-Mediated NIR Polymerization Amplification Strategy for Quantitative Determination of Lung Cancer-Related Nucleic AcidsAn electrochemical biosensor based on Cu₂（OH）PO₄-mediated near infrared light（near-infrared light,NIR）ATRP is built for the detection of lung cancer-related nucleic acidss in this work.ATRP initiator is ligated to PNA/DNA duplex by the coordination of phosphate-Zr⁴⁺-carboxylate.Under the irradiation of NIR,the Fenton-like of Cu₂（OH）PO₄ and H₂O₂ is activated to generate hydroxyl radical（HO^·）for initiating ATRP.Then,numerous monomers are modified on the electrode surface.The limit of detection is 2.6 f M（R²=0.993）,and the linear range is from 10 f M to 1000 p M.The minimum molar number of nucleic acids which can be detected is 26 zmol（10μL）.NIR has great potential in biological detection because of its strong penetration and little damage to biological tissues.（3）Photoinitiator I-2959-Induced Polymerization Amplification Strategy for Quantitative Determination of Lung Cancer-Related Nucleic AcidsAn electrochemical biosensor based on intramolecular photoinitiator（I-2959）-Induced ATRP is constructed to detect lung cancer-related nucleic acids with an ultrasensitive strategy.Under the irradiation of blue light,I-2959 can generate free radicals through the homolytic cleavage which can reduce Cu（II）to Cu（I）through the reduction of carbon free radicals,and polymerization is activated.A large number of electroactive probes are grafted from the electrode surface.Under optimal conditions,the limit of detection is 3.16 f M（R²=0.992）,and the linear range is from 10 f M to 1000 p M.The carbon free radicals generated by homolytic cleavage have little damage to biomolecules,which is more suitable for biomolecular detection.（4）EY-Mediated Metal-Free Photoinitiated ATRP Amplification Strategy for Quantitative Determination of Lung Cancer DNAAn electrochemical biosensor based on Eosin Y（EY）-mediated metal-free photoinitiated ATRP is constructed to achieve ultrasensitive detection of lung cancer DNA.Under blue light irradiation,the reduction quenching cycle is triggered with EY as the photocatalyst,and N,N,N’,N",N"-pentamethylethylenetriamine（PMDETA）is the electron donor.A large number of electroactive probes are grafted from the electrode surface controlled by reductive quenching cycle.Under optimal conditions,the limit of detection is 1.4 f M（R²=0.989）,and the linear range is from 0.1 p M to 10 n M.This strategy solves the problem of residual copper catalyst,and simplifies the construction process of biosensor.（5）g-C₃N₄-Mediated Photoinitiated RAFT Amplification Strategy for Quantitative Determination of Breast Cancer-Related Nucleic AcidsA novel biosensor based on g-C₃N₄-mediated photoinitiated reversible fragmentation chain transfer（RAFT）is constructed for the ultrasensitive detection of breast cancer-related nucleic acids.DNA tetrahedron is adopted as capture probe,which is fixed to the electrode surface by Au-S bond.Chain transfer agent of RAFT is linked to the electrode surface via sandwich structures between DNA tetrahedra,target DNA and detection DNA.Under the irradiation of blue light,g-C₃N₄ and tertiary amine-mediated RAFT is activated.Plenty of polymer chains are grafted from the electrode surface via chain transfer agent.The limit of detection is as low as 0.14 a M（R²=0.994）,and the linear range is from 10 a M to 1000 f M.This method also solves the problem of deoxygenation of cocktail and residual metal catalyst.