Study On Electrochemical Biosensors Based On Quantum Dot-labeled DNA Signal Amplification Technology

With the development of deoxyribonucleic acids(DNA)signal amplification technology,it has been widely used in basic biological research,biomedicine,food,etc.,and has become one of the most valuable analytical methods.In recent years,due to its advantages of simple operation and high sensitivity,DNA signal amplification technology has been widely used in basic biological research,biomedicine,biomolecule detection,etc.,and has become one of the most valuable analytical methods.At the same time,electrochemical analysis methods are often combined with DNA signal amplification technology for the detection of biological molecules(DNA,RNA,proteins,small molecules,etc.)due to their excellent small size,sensitivity,and low cost.This paper mainly constructs two kinds of detection of protein and m RNA based on DNA signal amplification.1.Electrochemical biosensor combined with immunoassay and enzyme activity analysis for accurate analysis of human apurinic/apyrimidinic endonuclease 1Human apurinic/apyrimidinic endonuclease 1(APE1)is an intracellular multifunctional enzyme that plays an important role in cell cycle control,transcription factor regulation,and oxidative stress response.In addition,APE1 also plays a key role in the base excision repair of nucleic acids.Here,we have developed a new electrochemical biosensing method that combines immunoassay and enzyme activity analysis for APE1 enzyme activity detection.The basic principle is to design and construct a DNA-catalyzed hairpin self-assembled sequence structure that can be activated by APE1,and mark the DNA of the structure with electrically active Cd S and Pb S quantum dots as a signal output molecule to output electrochemical signal.In this system,the generation of two positive signals need to meet both the conditions of immune recognition and enzyme activity catalysis,which is the basis for accurate enzyme activity analysis of APE1.Experimental results show that the linear range of this method for APE1 is 0.01-4 U/m L,and the detection limit is 0.00518 U/m L,which is equivalent to or better than conventional methods.More importantly,this method can accurately reflect the inhibitory effect of the inhibitor on the enzyme activity,and can also distinguish APE1 from its isoenzyme,effectively overcoming the defect that the isozyme cannot be distinguished only with DNA fluorescent probes.It also solves the problem that traditional ELISA immunoassay cannot accurately measure enzyme activity.The successful implementation of this integrated system will help the research and application of APE1 in the field of biomedicine,and also provide a reference for the accurate analysis of other enzyme activities.2.Study on the detection of miR208 a by polydopamine-assisted electrochemical biosensorAcute myocardial infarction(AMI)is myocardial necrosis caused by acute and persistent ischemia and hypoxia of coronary arteries.miR208 a is a target small RNA highly related to AMI.In this paper,we designed a hexagonal star nucleic acid signal amplifier(Hexagon Star G4,Hex SG4)containing G-quadruplex sequence and used it for electrochemical detection of miR208 a.The experimental principle is to integrate the G-quadruplex sequence with horseradish peroxidase(HRP)activity into three DNA hairpins(H1,H2,H3),which can pass through the complementary sequence forms a complete hexagonal star structure(Hex S).It is also possible to connect Hex S to magnetic beads with a capture nucleic acid strand through base complementation.In the presence of potassium ions and hemin,the G-quadruplex sequence in the hexagonal star structure can form a G-quadruplex and has catalytic activity,thereby oxidizing dopamine(Dopamine,DA)to form polydopamine(PDA).Then,the electrochemically active Cd Te Quantum dots(QDs)are precipitated by PDA,and finally the anode stripping voltammetry is used for electrochemical detection.After optimized design,the linear detection range of this electrochemical sensor for miR208 a is between 1 p M-10 n M,and the minimum detection limit is 0.3 p M.The electrochemical sensor is simple,and its sensitivity is much higher than that of fluorescence detection,which shows good application prospects in the detection of miRNA.