Construction Of Two Bio-based Electrochemical Sensing Platforms And Detection Of Disease Markers

Highly sensitive detection of disease-related biomarkers plays an important role in early diagnosis,treatment and drug screening.Related studies have shown that the changes of dopamine（DA）in human body fluid and microRNA（miRNA）in cancer cells are related to the occurrence of various brain nerve diseases and malignant tumors,respectively.Although a variety of hypersensitive biosensing platforms have been developed,there are still limitations on how to achieve the analysis and detection of biomarkers in complex actual samples.The development of new analysis strategies is still the focus of research.Electrochemical analysis has become one of the common methods for biomarker analysis in recent years due to its high sensitivity and easy operation.In this paper,different types of electrochemical sensing strategies for the detection of different biomarkers were constructed based on two biomaterials.Furthermore,a series of new strategies for electrochemical analysis of DA and cancer-related miRNA-21（miR-21）were established.The main research works are summarized as follows:（1）Using carbon dots（CDs）and laccase（Lc）as electrode modification materials,an enzyme-based electrochemical sensor has been constructed for the efficient detection of DA in human serum.The experimental results show that the introduction of CDs can effectively improve the electron transfer ability between the Lc and the electrode interface.At the same time,the presence of carboxyl（-COOH）,hydroxyl（-OH）,amino（-NH₂）and other groups on the surface of CDs is beneficial to DA.The specific adsorption at the electrode interface in turn increases the possibility of DA oxidation at the interface.Compared with traditional DA detection methods,the electrochemical sensor constructed in this study has good electrocatalytic activity,wide linear range（0.25μM-76.81μM）,low detection line（0.083μM）,anti-interference,repeatability and excellent stability.The recovery rate in human serum can reach 100.8%-103.9%.（2）In this study,by combining endonuclease-assisted target cycling with a primer exchange reaction（PER）cascade,a nuclease-promoted dual-cascade signal amplification electrochemical biosensing platform has been established for the detection of miR-21.miR-21 can be cyclically transcribed into a large number of single-stranded DNA fragments（s DNA）driven by endonuclease to complete the first amplification of miR-21signal.Subsequently,under the action of Klenow large fragment polymerase,the s DNA triggered the dumbbell probe（DP）,which can activate the PER cascade amplification process to generate a large number of nascent single-stranded DNA（cDNA）to complete the secondary amplification of the target signal.Using cDNA,DNA signaling molecule spheres loaded with a large number of electroactive molecules(Ru³⁺-DNS)can be captured on the electrode surface for highly sensitive electrochemical detection of miR-21.The electrochemical biosensor has a detection range of 0.2 aM-0.2 nM with a detection limit of 0.067 aM.The excellent selectivity enabled the biosensor to discriminate target miR-21 from other miRNAs,even miR-21 the single-base mismatched.Furthermore,the reliability of the method for specific detection of miRNAs in real samples was validated by analyzing the expression of miR-21 in human serum and cell lysates.