| Objective Parkinson’s disease(PD)is the second most common neurodegenerative disease after Alzheimer’s disease and is commonly seen in middle-aged and elderly people.There are about 7 million patients with Parkinson’s disease worldwide,and China is the most populous country in the world and is gradually entering an aging society.It is estimated that the number of PD patients in China will increase to 4.94 million by 2030,accounting for about half of the global PD patients,and the advanced symptoms of PD are very severe,which will impose a huge burden on the Chinese economy.The main reason for the appearance of symptoms in PD is the brain dopamine.As a major component of the neuropathological characteristic inclusions of PD,α-synuclein oligomers are neurotoxic and cytotoxic,and their abnormal aggregation suggests the pathogenesis and process of PD.α-synuclein oligomers are also considered as a biomarker of PD,and traditional detection methods such as ELISA are complicated and difficult to detect traces ofα-synuclein oligomers in plasma.The aim of this study is to establish an ultra-sensitive,low-cost and easy-to-use electrochemical biosensor for the detection ofα-synuclein oligomers in plasma and provide an idea or a method for clinical diagnosis and monitoring of PD.Methods In this study,a carbon-based OMC-MWCNTs-rGO-CS nanocomposites prepared from ordered mesoporous carbon(OMC),reduced grap Hene oxide(rGO),multi-walled carbon nanotubes(MWCNTs)and chitosan(CS)was used as modification material for the gold electrode,which has excellent electrical conductivity,and then the designed sensor was prepared by modifying the specific aptamer as a biorecognition component on the electrode surface.Electrochemical cyclic voltammetry(CV)was used to characterize the signal amplification and aptamer modification of the material;differential pulse voltammetry(DPV)was used to study the detection of the targetα-synuclein oligomers by the sensor with qualitative potential values and quantitative current values;in addition,experimental conditions such as the modification volume of OMC-MWCNTs-rGO-CS nanocomposites,concentration of aptamer,and incubation time of the aptamer and antigen were optimized in this study,and the sensor was verified under optimal conditions.The limit of detection,precision,accuracy,selectivity,reproducibility and stability of the method were explored under optimal conditions.Finally,the prepared sensors were applied to actual samples,i.e.,spiked human plasma samples,to evaluate their practicality.Results The OMC-MWCNTs-rGO-CS nanocomposites synthesized in this study had a significant electrochemical signal amplification.And the experimental optimal conditions were:the modification volume of OMC-MWCNTs-rGO-CS nanocomposites was 3μL,the concentration of aptamer was 10μmol/L,and the incubation time of aptamer antigen was 45 minutes.Under the optimal conditions,the logarithmic correlation between the current difference measured by DPV andα-synuclein oligomer concentration was good,and the equation of the standard curve was Y=1.610X+13.82,R~2 was 0.9956,and the linear range was 0.05 f mol/L～50 f mol/L.The peak potential of DPV was around0.20 V,and the value of peak current increased with theα-synuclein oligomers.The limits of detection(LODs)were 0.043 f mol/L according to the method specified by IUPAC.The precision(RSD)of the methods constructed in this study was less than 5%,and the spiked recoveries were between 75%and 105%.The interferences of Tau-441 protein(Tau-441),ascorbic acid(AA),glucose(Glu)and L-cysteine(L-Cys)were all less than 5%.The response values of different gold electrodes for the same concentration of samples were not significantly different,and the prepared sensors could maintain 92.69%of their initial signals after being stored at 4℃for one week.The constructed sensors were used again for the detection of spiked plasma samples,and their spiked recoveries were close to100%.Conclusion The electrochemical aptamer biosensor constructed in this study had successfully detectedα-synuclein oligomers in human plasma.The carbon-based OMC-MWCNTs-rGO-CS nanocomposites had excellent electrochemical signal enhancement and were suitable as modification materials for electrodes.The constructed sensor,using DPV technology,responded toα-synuclein oligomers in plasma in the linear range of 0.05 f mol/L to 50 f mol/L with a low detection limit of 0.043 f mol/L.The constructed sensor had good precision,accuracy,selectivity,reproducibility and stability.The constructed sensor method was used in actual plasma spiked samples and performed well,indicating the potential of the sensor for clinical diagnosis and monitoring of PD. |
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