Investigation And Application Of Electrochemical Aptamer And Protein Sensors Based On Three Carbon Nanomaterials

Electrochemical biosensor as an analysis device mainly composed of a recognition element and a transduction element.The recognizer consists of a sensitive biomolecule?such as aptamers and proteins?,which can high affinity and specificity recognize a target.The solid electrodes are usually selected as the transduction element,which can convert the generated biochemical signal into a measurable electrical signal.Electrochemical biosensors have the advantages of well-developed theory,facile design,convenient operation,low cost,and portability,which are widely used in the fields of biochemical technology,food industry,environmental analysis,biomedicine,and clinical diagnosis.With the development of nanotechnology and materials science,nano-biosensing technique has gradually entered the public vision with more and more kinds of nanomaterials used in the construction and investigation of biosensors.Among them,carbon nanomaterials are often selected as sensitizer for the modification of electrodes due to their high electrical conductivity,large specific surface area and good biocompatibility,which can bring with some advantages like achieving greater immobilization of biorecognition probes,enhancing the electrochemical performances and improving the detection sensitivity.This thesis focuses on the preparation and characterization of carbon nanofiber composites,construction of aptamer/protein-based sensors,and highly sensitive electrochemical detection of small molecules such as mercury ion(Hg²⁺),lead ion(Pb²⁺),trichloroacetic acid?TCA?,potassium bromate?KBrO₃?,sodium nitrite?NaNO₂?and so on,which can be summarized as follows:1.Polyacrylonitrile fiber?PANF?was prepared by electrospinning,and carbon nanofiber?CNF?was further obtained by carbonization under high temperature.Platinum nanoparticles?PtNPs?were loaded onto the surface of the CNF by hydrothermal synthesis to obtain a Pt@CNF nanocomposite.The composite was characterized by scanning electron microscopy.The result shown that the CNF had a spatially interlaced structure,and the PtNPs were successfully fixed on the fiber surface.Carbon ionic liquid electrode?CILE?was used as the basic electrode,and the prepared nanocomposite was modified on CILE surface,which was further covered with a layer of gold nanoparticles?AuNPs?by electrodeposition.The electrochemical performance of Au/Pt@CNF/CILE was tested with the result displayed it had good electrical conductivity and large specific surface area.According to the self-assembly method,aptamer probe?Aptamer?was immobilized on electrode,which could specifically recognize Hg²⁺.Therefore,an electrochemical aptamer sensor with high sensitivity,wide detection range,good reproducibility and stability was successfully constructed,as well as established an analytical method for efficiently and rapidly detecting Hg²⁺.2.CILE was used as the substrate for electrochemical transduction,which was covered with a layer of reduced graphene oxide?r GO?membrane by electroreduction method.Further,carboxylated multi-walled carbon nanotube?COOH-MWCNT?was modified on rGO/CILE surface,which could bring with unique electrochemical properties due to the synergistic effect of carbon materials.Then,AuNPs were formed on COOH-MWCNT/rGO/CILE interface using electrodeposition technique,which could provide specific binding site for the self-assembly of aptamer probe?Aptamer?through Au-S bond.Thioglycolic acid?TGA?was further pipetted on the sensing platform and used to eliminated the non-specific adsorption to obtain the electrochemical aptamer sensor?Aptamer/Au/COOH-MWCNT/rGO/CILE?.The immobilized aptamer probe could be induced to form G-quadruplex conformation by Pb²⁺,which was convenient for the insertion of Hemin.With Pb²⁺concentration increased and the electrochemical signal of hemin also increased,which could act as the indicator for the quantitative analysis of Pb²⁺.Therefore,this sensor was used to determine Pb²⁺with the detection range of 1.0×10^-13 mol/L to 1.0×10^-5 mol/L and the detection limit of 3.33×10^-14mol/L?3??.3.Based on the previous works,Au/Pt@CNF/CILE was selected as the sensing platform,and the capture probe?CP?was successfully assembled on the modified electrode surface by Au-S bond with TGA used to block the excess binding sites.The aptamer probe?AP?was further hybridized with CP to form a stable double-stranded DNA structure to construct an electrochemical aptamer sensor?AP/CP/Au/Pt@CNF/CILE?for the detection of Pb²⁺.Methylene blue?MB?was used as indicator,which was embedded into the double strands.Since Pb²⁺could induce G-rich DNA to form allosteric G-quadruplex oligonucleotides,which resulted MB released from the double strands.Therefore,the quantitative detection of Pb²⁺was realized by collecting the electrochemical responses of MB before and after incubated with Pb²⁺.The constructed aptamer sensor displayed better sensitivity and selectivity for Pb²⁺detection with the concentration range of 1.0×10^-16 mol/L to 1.0×10^-11 mol/L and the detection limit of 3.3×10^-1717 mol/L?3??.4.Nano cobalt oxide?Co₃O₄?was mixed with polyacrylonitrile?PAN?to prepare Co₃O₄doped CNF nanocomposite?Co₃O₄-CNF?by electrospinning and carbonization techniques,which was further used to modify on CILE surface.Hemoglobin?Hb?was immobilized on Co₃O₄-CNF/CILE surface with Nafion acted as the protective film to fabricate an electrochemical protein biosensor?Nafion/Hb/Co₃O₄-CNF/CILE?.Electrochemical behavior of Hb on modified electrode was investigated with a pair of quasi-reversible redox peak appeared on cyclic voltammogram and relevant electrochemical parameters were calculated.Moreover,this biosensor had good analytical capabilities for electrocatalytic reduction of different substrates including TCA,KBrO₃ and NaNO₂ with wide detection range from 40.0to 260.0 mmol/L,0.1 to 48.0 mmol/L and 1.0 to 12.0 mmol/L,respectively.The prepared sensor was also used for the quantitative detection of real samples,which displayed the great practical ability.