Research On Electrochemiluminescence Biosensor Based On Novel Nanomaterials And Multiple Signal Amplification Strategies

The heavy metal ions in the environment seriously affect the growth of organisms,and enter the human body through enrichment to endanger the nervous and renal systems.Biomarkers are usually used to diagnose diseases,assess or predict the efficiency and safety of drugs.Electrochemiluminescence(ECL)technology is widely used in biomolecular detection,food and drug analysis and environmental monitoring due to its low background interference,high reproducibility and accuracy.At the same time,people also actively explore new nanomaterials and use a variety of signal amplification strategies,such as hybrid chain reaction,rolling ring amplification reaction and so on to greatly reduce the detection limit of the target.In this paper,a variety of new nanomaterials have been used and coupled with many signal amplification strategies to successfully construct electrochemiluminescence biosensors with good performance for the highly sensitive detection of Pb²⁺,PSA,and Let-7a micro RNA.This article mainly performed the exploration from the following two aspects:1.In this work,a new DNA nanotweezers-Ag nanoparticle(NP)as signal-switching probe is designed and coupled with tripod-like capture DNA structure to construct an ultrasensitive“ON-OFF”electrochemiluminescence(ECL)biosensor for detecting Pb²⁺.The unique tweezer nanostructure has two G-base-rich sequences on different arms,and the G4-chain structure is simply constructed by switching tweezer conformation from“open”to“close”,which can be controlled by alternative target released in Pb²⁺triggered cyclic amplification process.The activated G4-structure can specifically capture plenty of hemin to the closed state,which has a strong quenching effect on ECL of Ru(phen)₃²⁺.After reduced graphene oxide(ERGO)with good electrochemical activity and loading capacity is electrodeposited on the electrode,Ru(phen)₃²⁺as ECL probe is strongly adsorbed on it via?-?conjugation.Then tripod-like DNA structure with spiral extensibility and no backfilling was linked to the surface to capture hemin/G4-structure,resulting in significant decrease of ECL for sensitive Pb²⁺detection.Due to the design flexibility of DNA nanotweezers structure,the strategy provides universal method of ECL biosensors for a variety of metal ions and other bioassays.2.In this work,a versatile Au nanoclusters/Au-MnO₂nanoflowers electrochemiluminescence(ECL)resonance energy transfer(RET)platform coupled with rolling circle amplification technique was developed for“OFF-ON”detection of prostate antigens(PSA)and Let-7a miRNA(a cancer biomarker).The prepared methionine-stabilized Au nanoclusters(Met-AuNCs)displayed dramatically enhanced ECL intensitity and stability by using 3,4-thiophene dicarboxylic acid modified electrode.Efficient ECL-RET system between Met-AuNCs donors and Au-MnO₂nanoflowers(NFs)acceptors was constructed and used to develop a sandwich ECL immunosensor for signal“OFF”detection of PSA.Furthermore,the second target Let-7a miRNA triggered rolling circle amplification(RCA)reaction to produce long DNA nanowires,which can bridge a large number of biotin-streptavidin structure to label plenty of alkaline phosphatase(ALP),and ALP catalyze L-ascorbic acid-2-trisodium phosphate(AAP)to produce abundant ascorbic acid(AA).Then the product AA in-situ reduced(etched)MnO₂ nanoflowers(NFs)to Mn²⁺,which inhibited ECL quenching of MnO₂ NFs to Met-AuNCs,and recovered ECL to significantly high“ON”signal for Let-7a detection.Importantly,this multi-functional versatile ECL platform combines advantages of 3,4-thiophene dicarboxylic acid-enhanced ECL of AuNCs,high loading and quenching ability of Au-MnO₂ NFs,target-initiated RCA,enzyme-catalyzed reaction for assay of double targets.The proposed multifunctional biosensing system is also suitable for highly sensitive detection of other disease-related proteins with promising actual and clinical application.