Application Of Dual-signal Electrochemiluminescence Sensing Strategy For Detection Of Myocardial Injury Markers

Electrochemiluminescence（ECL）is a novel analysis technology combining the electrochemical and chemiluminescent.With the advantages such as simplified optical setup,low background signal,high sensitivity and fast analysis speed,it has been widely applied in clinical medicine,food,medicine,environmental monitoring and other fields.ECL technology can not only realize the detection of the single target,but also can realize the simultaneous detection of multiple targets.Single-target detection based on single-signal is susceptible to environmental and instrumental disturbances,which may lead to false positive or false negative results.The dual-signal ECL ratiometric strategy for single-target detection could effectively eliminate external interference,achieving accurate detection of the target,thus arousing widespread concern.However,for clinical analysis,the detection of a single biomarker is usually not enough to achieve an accurate diagnosis of the disease.The multiplexed targets detection could significantly improve the diagnostic accuracy and efficiency,showing great applications in clinical diagnosis.Acute myocardial infarction（AMI）as a usual human health problem,has become the most common cause of death in the past decades.Studies have shown that the levels of brain natriuretic peptide（BNP）,cardiac troponin I（c Tn I）and micro RNA（mi RNA）will change in early myocardial injury,thus they are regarded as markers of myocardial injury.It is of great significance to realize the sensitive and accurate quantitative detection of myocardial injury markers,and could effectively reduce the risk of death.In this paper,three kinds of biosensors were constructed based on the dual-signal for dual target combined detection and dual-signal for single target ratio detection modes to realize the highly sensitive detection of different myocardial injury markers.The main researchs are included as follows:（1）Potential-resolved ECL multiple immunoassay for myocardial injury markers BNP and cTnIIn this paper,a potential-resolved ECL immunosensor was constructed,using graphiticphase carbon nitride nanosheet（g-C₃N₄）and N-（aminobutyl）-N-（ethylisoluminol）（ABEI）as cathodic and anodic emitters,respectively,to realize the simultaneous detection of the dual target BNP and c Tn I.Firstly,Au nanoparticles were modified onto the electrode surface by electrodeposition method to capture the anti-BNP₁and anti-c Tn I₁.The antigen of BNP and c Tn I,secondary antibody composites anti-BNP₂-Au NPs@g-C₃N₄ and anti-c Tn I₂-Au NPs@ABEI were stepwise incubated onto the electrode surface.With the increase in the concentration of BNP and c Tn I,the ECL signals at+0.7 V from ABEI and-1.5 V from g-C₃N₄ were simultaneously increased,thus achieving the synchronous immunoassay of BNP and c Tn I.The integration of ABEI/O₂ and g-C₃N₄/S₂O₈^2-system avoided the defects of the currently multiplexed immunoassay,thus providing an attractive ECL platform for synchronous determination of multiple biomarkers at the same sensitive interface.（2）A ratiometric electrochemiluminescence biosensor based on BP-Cd Te QDs with dual-emission for detecting mi RNA-126The traditional ECL ratiometric assay is usually based on two different ECL emitters,and the choice of two suitable emitters and shared co-reagents makes this strategy challenging to construct.The single-emitter-based ratio ECL model could overcome the limitation of traditional ECL ratiometric assay and simplify the construction process.In this work,Cd Te quantum dots equipped with black phosphorus nanosheets（BP-Cd Te QDs）were developed to simultaneously emit the cathodic and anodic ECL signals,thus were used as a single-emitter,which combined with the cascade nucleic acid amplification strategy to construct ratio ECL strategy for mi RNA-126 detection.Through the target recycling triggering rolling-circle amplification（RCA）reaction,a large number of glucose oxidase modified single-strand 1 was introduced to in situ generate H₂O₂,which acted as a dual-role moderator to quench the anodic ECL emission while enhance the cathodic emission,thereby realizing the sensitive detection of mi RNA-126 with the detection limit of 29 a M（S/N=3）.This is the first time that the cathodic and anodic ECL emissions of Cd Te QDs have been developed for the ratio analysis.The combination of dual-ECL-emitting BP-Cd Te QDs with dual-regulating H₂O₂expanded the application of Cd Te QDs in the ECL field and provided an attractive ECL ratio platform for biological analysis.（3）A ratiometric ECL biosensor based on SnS₂ quantum dots hollow porous polymeric nanospheres and Ir nanorods for the sensitive detection of cTnIIn this work,SnS₂ quantum dots（SnS₂ QDs）hollow porous polymeric nanospheres（SnS₂-HPNSs）and Ir nanorods（Ir NDs）were respectively used as cathodic and anodic ECL emitters to construct the potential-resolved ratiometric strategy for the sensitive detection of c Tn I.Firstly,SnS₂-HPNSs were synthesized by layer-by-layer assembly of oppositely charged moiety polymers,and then combined with gold nanoparticles,norepinephrine（NA）and anti-c Tn I₂（Ab₂）to prepare secondary antibody complex Ab₂/NA/Au NPs/SnS₂-HPNSs.Then,the Ir NDs were modified onto the electrode surface,which were used as the matrix for immobilizing the anti-c Tn I₁（Ab₁）,and further capturing the antigen c Tn I and the secondary antibody complex Ab₂/NA/Au NPs/SnS₂-HPNSs via the immune reaction.With the increase of the target c Tn I,the NA and SnS₂-HPNSs introduced by the secondary antibody complex increased,and the cathodic signal from SnS₂-HPNSs would gradually increase.Meanwhile,the anodic signal from Ir NDS would gradually decrease since NA can effectively quench the ECL emission of Ir NDS.Thus a highly sensitive detection of c Tn I was achieved.Significantly,HPNSs can not only achieve the high loading of SnS₂ QDs at the three-dimensional level,but also effectively reduce the inner filter effect of SnS₂ QDs,thus obviously improving the ECL efficiency of SnS₂-HPNSs.The combination of SnS₂-HPNSs and Ir NDs provided a promising ECL ratio platform and a new strategy for biological analysis.