Technology And Application Of Electrochemiluminescence Based On Signal Amplification Strategy

Electrochemiluminescence(ECL)has become one of the most important bioanalytical detection methods due to its powerful analytical capabilities.Electrochemiluminescence initiates and controls electrochemical reactions to generate electronic excited states by applying electric potential.It is essentially a chemiluminescence reaction on the electrode surface.Since the early detection of various cancer markers and the diagnosis of diseases require ultra-sensitive and high-precision bioassays,the development of ECL as a measurement technology has been promoted.The signal amplification strategy is introduced into the ECL field,and combined with ECL active substances for sensitive detection of the target,it has been studied to obtain better performance and lower detection limit.In this thesis,biosensors were designed to detect Zika virus RNA and VEGF₁₆₅protein through enzyme-assisted nucleic acid amplification and peroxidase catalysis,and the ECL of single particles was studied by laser thermal induction.Imaging method.The main research contents are as follows:1?Metal-organic gel and metal-organic framework based switchable electrochemiluminescence RNA sensing platform for Zika virusIn this work,an ultrasensitive switchable electrochemiluminescence(ECL)RNA sensing platform was established based on metal-organic gel(MOG)and metal-organic framework(MOF)as electrode matrix and nanotag,respectively.The former was prepared via the in-situ loading of graphite-like carbon nitride(g-C₃N₄)and post-assembly of Au nanoparticles(AuNPs)within Zr-based MOG(AuNPs&g-C₃N₄@Zr-MOG),exhibiting highly efficient solid-state ECL.Fe-MIL-88 MOFs not only act as an ECL acceptor in ECL resonance energy transfer system but also have metal active centers to consume co-reactant,initiating double quenching effect on the ECL of AuNPs&g-C₃N₄@Zr-MOG.Then,DNA probe containing an apurinic/apyrimidinic(AP)site connected the both,resulting in a turn-off signal.In the presence of the target RNA,the ECL was turned on because the AP site of DNA probe was activated to be circularly cleaved by endonuclease IV.Using Zika virus(ZIKV)RNA as a model analyte,the ECL platform showed a wide detection range from 0.3 n M to 3?M with a detection limit as low as 0.1 n M.Besides,the sensor exhibited excellent specificity and stability,as well as acceptable real sample detection capability,illustrating the feasibility of clinical measurement in the future.2?Fe(?)-Organic Gels Substrate-Catalyzed Dual Electrochemiluminescence Signal Systems for Direct Evaluation of the VEGF₁₆₅ Expression on Cells SurfaceThis work established a new proportional electrochemiluminescence cytosensor platform catalyzed by Fe(?)-organic gels matrix.Luminol-capped Au nanoparticles(AuNPs-luminol)were embedded in situ in Fe-MOG and served as a sensing platform for cell capture at anode response.At the same time,due to the high impedance of the cells,the ECL signal was quantitatively suppressed,resulting in a negative correlation between the anode ECL signal of luminol and the number of captured cells.At the same time,aptamer-modified CdS QDs(CdS apt)was usd as the cathodic response,targeting on the cell surface to indicate the expression of VEGF₁₆₅ protein.In addition,the MOG with highly dispersed Fe(?)catalytic centers in the composite material could be used as a co-reaction accelerator to generate a large amount of ROS to enhance the ECL signal at both two potentials.The detection method developed by us shows excellent performance in analyzing the level of VEGF₁₆₅.This strategy can also be used to evaluate the content of VEGF₁₆₅ in different cell systems and further extended to clinical testing.3?Laser-induced thermal convention ECL microscopy of enhancing ECL intensityElectrochemiluminescence imaging has become an indispensable branch of microscopic imaging technology due to its advantages of good temporal and spatial resolution and high throughput.However,it is still difficult to apply in the field of bioimaging in terms of single particles'weak light intensity and high luminescence potential.Based on this,we independently built a set of equipment that combined a 730 nm laser with an electrochemiluminescence microscope to observe the effect of the laser on the ECL signal.In the work,by observing the ECL intensity changes before and after the laser is applied on the side and front,it is found that the laser can effectively enhance the ECL intensity,and the proposed mechanism is further proved by the naphthol obstructing convection and fluorescence observation.Finally,we used this enhancement technology to achieve ECL enhancement of gold nanoparticles and RuDSN.Based on the principle that the thermal convection induced by laser is used to enhance ECL signal,it has good repeatability and operability,which paves the way for the application of single particles to ECL organelle imaging.