Development Of Nanomaterial Electrochemical Biosensors For Detection Of Neurodegenerative Diseases Biomarkers

Neurodegenerative diseases are a kind of chronic progressive brain diseases with dormant onset and irreversible,which endanger the health,safety and self-care ability of the elderly.Alzheimer’s disease（AD）and Parkinson’s disease（PD）are the most widely distributed and influential neurodegenerative diseases in the world,and there is no effective treatment in the world.If AD and PD can be screened in the early stage,the health safety and quality of life of the elderly can be guaranteed,and the social and economic burden caused by the disease can be reduced.The natural forms ofβ-amyloid（Aβ）,tau protein andα-synuclein（α-syn）participate in the composition of neurons in the central nervous system,perform their functions normally and maintain the normal life function of the human.When the human body is in a pathologic state,these proteins misfold and form toxic polymers（oligomers and fibrils）.As the accumulation of polymer exceeds that of clearance,the body will undergo oxidative stress response,which kills the neurons cells,and eventually leads to the occurrence of AD and PD.At present,the enzyme-linked immunosorbent assay（ELISA）,immunoblotting,immunohistochemistry,liquid chip and surface plasmon resonance（SPR）for detecting Aβoligomer（Aβo）,tau381 protein andα-syn oligomer（α-syno）are generally sensitive and time-consuming,and ELISA kit is relatively expensive.With the increasing aging,the incidence rate of AD and PD is increasing year by year.It is necessary to develop specific,sensitive,rapid and economical detection techniques.Electrochemical biosensing technology has become a research hotspot of protein detection technology because of its advantages of high sensitivity,rapid response,simple operation and low cost.In recent years,nanomaterials,due to their size in the nanoscale,have different properties from macromaterials,and they have been widely used in information industry,energy and environmental protection,sensors,biomedicine and other fields.Carbon nanomaterials and gold nanomaterials have the advantages of controllable size,high specific surface area and excellent electronic properties.Nanocomposites can be synthesized by one-step or two-step method and combined on the electrode surface to improve sensitivity.However,the simple glassy carbon electrode（GCE）cannot specifically recognize proteins,and its electron transfer efficiency was very low.Researchers have found that aptamers have similar specificity to antibodies,and they have strict recognition ability and high affinity for binding target.Therefore,electrochemical biosensors with high sensitivity and specificit were prepared for the detection of AD biomarker（Aβo and tau381）and PD biomarker（α-syno）using nanocomposites as signal amplification materials and antibody and aptamer as recognition elements in this paper.The four researches are described as follows:1.Tau381 electrochemical aptasensor was successfully prepared by using CG/TH/Au NPs nanocomposites synthesized from carboxyl fossil ink（CG）,thionine（TH）and gold nanoparticles（Au NPs）.Tau381 was quantitatively detected by differential pulse voltammetry（DPV）using TH as redox probe.The morphology of CG/TH/Au NPs nanocomposites were characterized by field emission scanning electron microscopy.Electrochemical impedance spectroscopy（EIS）and cyclic voltammetry（CV）were used to study the detection performance of tau381 with different modified electrodes.The results showed that the impedance value（or current value）of CG/TH/Au NPs-GCE was significantly less than（or greater than）CG-GCE and CG/TH-GCE.Moreover,there was an excellent linear relationship（R²=0.9970）between the current difference of DPV and the logarithm of tau381 concentration in the concentration range of 1.00 p M-100.00 p M.The detection limit was 0.70 p M.In addition,the aptasensor was proved to have good reproducibility and specificity,and maintained good stability within 14 days,the recovery range was 97.90%-101.80%.And it was initially applied to blood samples.2.TH is a positively charged planar aromatic molecule,which can form a wideπ-πconjugated structure with carbon nanomaterials（r GO and MWCNTs）.TH/r GO/MWCNTs nanocomposite was synthesized by one-step hydrothermal method to modify GCE.A novel electrochemical aptasensor with Fe[（CN）₆]^3-/4-as redox probe was prepared for DPV detection of Aβo.The morphology of TH/r GO/MWCNTs nanocomposite was characterized by transmission electron microscopy（TEM）.In sulfuric acid and potassium ferricyanide/potassium ferricyanide solutions,TH/r GO/MWCNT-GCE has a stronger CV response than TH/r GO-GCE.The excellent linear relationship（R²=0.9942）between the current difference and the logarithm of Aβo concentration（0.0443 p M to 443.00 p M）,and the detection limit was 0.01 p M.The Aβo aptasensor showed good specificity and reproducibility,and maintained good stability within 15 days.The recoveries ranged from99.77%to 103.84%.The Aβo aptasensor was initially applied to blood samples.3.Poly（D-glucosamine）（PDG）has good biocompatibility,film stability,thermal stability,biodegradability and non-toxic.PDG/Au NPs/r GO/MWCNTs nanocomposites were synthesized by one-step method and modified on the surface of GCE.An ultra sensitive electrochemical immunosensor was successfully prepared using antibody as recognition element.square wave voltammetry（SWV）was used to detecteα-syno.The morphology of the nanomaterials was characterized by TEM,and the detection performance ofα-syno with different modified electrodes was recorded by CV and EIS.Due to the synergy of Au NPs,MWCNTs and r GO,α-syno electrochemical biosensor exhibited excellent electronic properties,fast electron transfer rate,excellent sensitivity,and specificity.In the range of 0.05 f M to 500 f M,there was a good linear relationship（R²=0.9936）between SWV current difference and logarithm of concentration,and the detection limit was 0.03 f M.α-syno immunosensor maintained good stability within 14days,the recovery ranged from 97.66%to 100.95%,and it has good reproducibility and specificity.4.Poly（thionine）（p TH）was modified on the GCE surface by electropolymerization,and gold nanoparticles（Au NSs）synthesized by N-2-hydroxyethyl piperazine-N’-2-ethanesulfonic acid were modified on the surface of p TH-GCE,and p TH/Au NPs nanocomposites were synthesized step by step to prepareα-syno electrochemical aptasensor.The morphology of the nanocomposites was characterized by TEM,and the electrochemical properties ofα-syno with modified electrodes were studied by CV and EIS.Theα-syno concentrations were from 0.1 a M to 10.00 f M and detection limit was0.07a M by EIS.Compared with p TH-GCE,Au NSs-GCE,TH-GCE and Au NPs-GCE,the response of p TH/Au NPs-GCE was the most obvious.Due to the fast electron transfer ability and large specific surface area of p TH and Au NSs,the sensitivity could be improved and the detection limit could be reduced.α-syno aptasensor maintained good stability within 14 days,and the recovery ranged from 98.41%to 101.77%,and it displayed good specificity and reproducibility.It showed good reliability in heparin and ethylenediamine tetraacetic acid（EDTA）anticoagulant plasma.The nanocomposites accelerate the electron transfer rate,and antibody and aptamer as recognition element improved the specificity of the electrode.The electrochemical biosensor with high sensitivity and strong specificity was prepared for the detection of trace Aβo,tau381 protein andα-syno in blood,providing new methods and new ideas for the early detection of AD and PD.It is of great significance to promote the health safety and quality of life of the elderly.