Study On Electrochemical Aptasensor Based On Novel Nanoflower Composites

Electrochemical biosensor is a highly sensitive detection technique,which is developed by combining biometric elements with electrochemical analysis.In recent years,it has been widely used in biochemical analysis and clinical diagnosis because of its low cost and simple instrument.To improve the analytical performance of electrochemical biosensor and realize sensitive detection of various tumor markers are of great significance to promote early accurate diagnosis of related tumor diseases.It is also a hot topic in current analytical chemistry research.In this thesis,three kinds of hybrid nanoflowers composites were synthesized by changing the organic ingredient during the self-assemble process,including HRP-Cu₃（PO₄）₂HNF,aptamer-Cu₃（PO₄）₂HNF&GO and SA-Cu₃（PO₄）₂HNF&GO.Aptasensors were constructed using them as nanocarriers and sensing substrate for detection of biomarkers.The specific work was as follows:1.A sandwich-type electrochemical aptasensor via biocatalytic precipitation amplification and by using gold nanoparticle compositesA sandwich-type electrochemical aptasensor is described based on two kinds of nanomaterials for detecting the carcinoembryonic antigen（CEA）with high sensitivity.The first was obtained by modifying gold nanoparticles with reduced graphene oxide and hemin（Hemin-r GO-Au NPs）.The second consists of HRP-Cu₃（PO₄）₂ hybrid nanoflowers linked to gold nanoparticles to obtain an architecture of type（HRP-Cu₃（PO₄）₂-HNF-Au NP）.They serve as carriers for two aptamers（apt1 and apt2）against CEA.Simultaneously,they were used to catalyze the precipitation reaction between 4-chloro-1-naphthol（4-CN）and H₂O₂.A sandwich-type assay linked to enzyme inhibition amplification was established for electrochemical determination of CEA.Under optimal experimental conditions,the response currents have a good linear relationship with the logarithm of CEA concentration in the range of 100 fg m L^-1 and 100 ng m L^-1.The detection limit was obtained as low as 29 fg m L^-1.2.Electrochemical aptasensor based on aptamer-Cu₃（PO₄）₂HNF&graphene oxideA signal-off aptasensor was described based on aptamer-Cu₃（PO₄）₂ hybrid nanoflowers&graphene oxide（aptamer-Cu₃（PO₄）₂HNF&GO）as an effective sensing platform and molybdophosphate precipitation as the signaling element.The aptamer,as the biorecognition unit grafted on aptamer-Cu₃（PO₄）₂HNF&GO by the specificity of streptavidin and biotin,captured the CEA through specific interaction.The molybdophosphate precipitation was formed by the reaction of the aptamer-Cu₃（PO₄）₂HNF&GO with molybdate.When aptamer-Cu₃（PO₄）₂HNF&GO specifically recognized CEA,the CEA-aptamer bioconjugate layer films not only hinder electron transfer on the sensing interface,but also impede the formation of molybdophosphate precipitation,resulting that the current signal significantly decreased.Under optimal experimental conditions,the response currents of the aptasensor have a good linear relationship with the logarithm of CEA concentration in the range of 10 fg m L^-1to 500 ng m L^-1.At the same time,the sensing also showed satisfactory results in the analysis of clinical serum samples.3.Electrochemical aptasensor based on target-triggered allosteric switching of aptamer beacon and nanoflowers&graphene compositesA label-free electrochemical aptasensor was developed based on the streptavidin-inorganic hybrid nanoflowers decorated with graphene composites（SFG）for the sensitive detection of interferon-gamma（IFN-γ）.To do this,a diblock dual-aptamer allosteric hairpin（DDAH）was designed,followed by conjugation with Au NPs（DDAH&Au NP）.The presence of target destroyed the stable hairpin structure,and then the catalytic cleavage of DNAzymes removed the IFN-γ-binding molecules,triggering the allosteric switching from inactive hairpin to active streptavidin aptamer（A-DDAH&Au NP）.Moreover,SFG were synthesized and used as substrates to modify glassy carbon electrodes.SFG specifically bind to the A-DDAH&Au NP to realize high-efficient readout of signals.Under the optimal conditions and by using differential pulse stripping voltammetry（DPSV）,the response currents have a good linear relationship with the logarithm of IFN-γconcentration in the range of 100 fg m L^-1 and 500 ng m L^-1.The detection limit is as low as 19 fg m L^-1.