Research On Electrochemiluminescence Biosensors Based On Novel Nano-emitters And Multiple Signal Amplification Strategies

Electrochemiluminescence?ECL?analysis combines the high sensitivity of chemiluminescence analysis with the high controllability of electrochemical analysis,which has become one of the frontier areas of modern analytical science.It is especially suitable for the analysis of biological systems with extremely low concentrations and complex components.The improvement of analysis sensitivity has been the research subject in the ECL biosensing field,which promoted the development of a series of new concepts and technologies for signal amplification.However,the development of an ultra-sensitive ECL biosensor still faces the following two challenges:?1?How to overcome the defect that the ECL efficiency of nano-emitters are lower than that of classical ECL reagents,and it is of critical importance to explore new ways to significantly improve ECL efficiency of nano-emitters.?2?How to improve the specificity and efficiency of recognition elements,and provide new strategies for high-selection and high-sensitivity detection of targets.Therefore,this paper mainly uses new ternary ECL systems to improve the ECL efficiency of nano-emitters,combined nucleic acid amplification or DNA nanomachine amplification strategies,constructs a variety of ECL biosensors to achieve ultra-sensitive detection of targets,providing new ideas for ECL bioanalysis.The research work of this paper is mainly divided into the following parts:1.Silver ions as novel coreaction accelerator for remarkably enhanced electrochemiluminescence in a PTCA/S₂O₈^2-system and its application in bioanalysis.In general,organic perylene compounds can be prepared into different nanostructures by supramolecular interactions,such as flake,sphere,block and line.However,the ECL efficiency of perylene compounds is limited by using peroxydisulfate(S₂O₈^2-)as coreactant,and it is necessary to explore new ways to improve their ECL efficiency.In this study,the metal ion of Ag?I?ion was used as a co-reaction accelerator to achieve strong and stable ECL emission of3,4,9,10-perylenetetracarboxylic acid?PTCA?supramolecular nanorod in a low-concentration S₂O₈^2-solution.Specifically,Ag?I?ion could react with S₂O₈^2-to produce strongly oxidizing intermediate(SO₄^·-)and Ag?II?ion.Then,the accompanying Ag?II?ion reacted with H₂O to generate more reactive intermediate species?ie.,hydroxyl radical?,which further accelerate the reduction of S₂O₈^2-to produce more SO₄^·-.In addition,the recycle of Ag?I?ion and Ag?II?ion could be achieved through electrochemical reactions.Therefore,an avalanche-type reaction was triggered to generate a large amount of SO₄^·-,which could react with PTCA to generate more excited states of PTCA,achieving a strong ECL signal.Based on the Ag?I?ion as an effective co-reaction accelerator in PTCA/S₂O₈^2-/Ag?I?ion system and dual nucleic acid amplification strategy,a new ECL biosensor was constructed for ultra-sensitive detection of mercury ions with the linear range from 1 fmol/L to 100 pmol/L,and the detection limit down to 0.33 fmol/L.This work firstly utilized metal ions as coreaction accelerator and provided a promising approach to improve the sensitivity of target analysis in ECL biosensing fields.2.SnS₂ quantum dots as new emitters with strong electrochemiluminescence and its application in bioanalysis.At present,many ECL quantum dots?QDs?,such as CdTe,PbS,and CsPbI₃,exhibited inevitable biotoxicity due to the leakage of toxic metal ions,which limited their application in the biosensing field.Therefore,the exploration of new QDs as ECL emitters with excellent biocompatibility is a challenging and interesting subject.In this study,a biocompatible,water-soluble and stable SnS₂ QDs was synthetized by a L-cysteine-assisted hydrothermal method?“down-top”approach?,which could act as a strong ECL nanoemitter in S₂O₈^2-solution.Furthermore,the smart circular peptide-DNA nanomachine was firstly designed,which could initiate the subsequent cascade recycling amplification to output massive mimic target products.Based on the SnS₂ QDs as ECL emitter and the 3D hierarchical silver nanoflowers as a co-reaction accelerator,a highly sensitive ECL biosensor was designed for antibody detection via smart circular peptide-DNA nanomachine amplification.The detection range was from1 fmol/L to 100 nmol/L,and the low detection limit was 0.33 fmol/L.Importantly,this work not only utilized SnS₂ QDs as a promising ECL emitter for biosensor construction,but also opened up an effective approach for sensitive immunological detection of nucleic acid cycle amplification technology.3.Pore confinement-enhanced electrochemiluminescence on SnO₂ nanocrystal xerogel with NO₃^-as co-reactant and its application in analysisMetal oxide nanocrystals are emerging ECL emitters with good biocompatibility and low toxicity,but their low luminescent efficiency is a critical drawback.Inspired by“pore confinement effect”of many porous nanomaterials with a unique,confined environment,the porous nanomaterials can open a new way to improve the ECL efficiency of weak emitters.In this study,a non-toxic and low-cost SnO₂ NCs was used to synthesize the porous SnO₂ NC xerogel,which exhibited the pore confinement-induced ECL enhancement behavior with NO₃^-as a co-reactant.In view of the fact that NO₃^-could be easily reduced to active nitrogen species for nanocrystal-based ECL amplification with the excellent stability and good biocompatibility.Therefore,on the basis of the new SnO₂ NC xerogel/NO₃^-system,a novel“signal-off”ECL sensing platform was developed for SO₃^2-detection with with the linear range from 1 pmol/L to 10 nmol/L,and the detection limit down to 0.33pmol/L.This work presented a new strategy of pore confinement-induced ECL enhancement that opened a new avenue to amplify the ECL signal from metal oxide NC.4.Analogous proximity-induced split G-Quadruplex as a new-generation of DNA nanodevice and its application in bioanalysisMost of the dynamic DNA nanodevices depend on the operation with the toehold-mediated strand-displacement?TSD?reaction.There is a dilemma between their stability and reactivity:either with a high-energy barrier to minimize leakage and improve stability or with a low-energy barrier to maximize catalysis and increase reaction activity.Herein,a split G-quadruplex?GQ?nanodevice consisting of four metastable dual?-DNA units was designed,which exhibited unprecedented advantages over the traditional duplex DNA nanodevice.First,four metastable dual?-DNA units were joined together to form a stable and rigid split GQ nanodevice by the analogous proximity effect,thereby minimized network leakage and improved molecular recognition.Second,this split GQ nanodevice could provide the high localized concentration of component,the excellent accessibility of target molecule,and the toehold binding region in an overhang domain to make the short down/top arm easily dehybridized,thus achieving fast TSD kinetics.As a proof of concept,the split GQ nanodevice-based ECL biosensor was successfully applied for antibody detection with the linear range from 1 fmol/L to 10 nmol/L,and the detection limit down to 0.32fmol/L.Significantly,this work gave impetus to the design of new generations of dynamic DNA nanodevices beyond the traditional ones with potential applications in sensing analysis,molecular imaging,and drug delivery.5.Matrix-induced electrochemiluminescence enhancement of hydrogel-confined gold nanoclusters and its application in analysisMetal nanoclusters?Au NCs?have become attractive ECL nanoemitters due to their fascinating molecular-like physicochemical properties.However,there are few reports on the relationship between Au NCs and microenvironment in ECL process.In view of this,a facile strategy has been developed to in situ synthesize a new class of glutathione-stabilized Au NCs confined in hydrogel matrix,which exhibited the highest ECL efficiency(?_ECL,up to 94.8%).This booster ECL phenomenon was rationally ascribed to the matrix-induced ECL enhancement modulated by the increased electron-hole recombination efficiency and the suppressed nonradiative relaxation.Benefiting from these advantages,the strong ECL emission was firstly observed in coreactant path,which located at⁷42 nm corresponding to surface-state transition.As a proof of concept,a rapid and sensitive sensing platform was successfully applied for S^2-detection by effectively quenching of S^2-to Au NCs/triethylamine system with the linear range from 0.1 pmol/L¹00 nmol/L and the detection limit down to 32fmol/L.This work extends our mechanistic understanding of the relationships of luminophores and microenvironment and applies this understanding to develop a high-performance ECL material in sensing and imaging field.