Study On New Methods In Photoelectrochemical Biomarker Detection

As an evolutionary generation of electrochemical bioassay,photoelectrochemical(PEC)bioanalysis inherited its advantages of rapid respond and low background,meanwhile,due to the different energy form of the excitation source and detection signal,PEC technique possessed higher sensitivity and demonstrated simple instrument and low cost compared to optical methods,thus becoming a hot topic among the community.Especially,much effort has been exerted in developing innovative recognition and signaling strategies according to its unique feature,and various biomarker,such as disease-related nucleic acids,proteins and small molecules,has been determined aiming at reliable clinical diagnosis.However,as a newly appeared analytical method,it's still inadequate in accuracy and practicability for clinical diagnosis and point of care application.Based on the state of art PEC biomarker detection and its future development,the following four protocols has been established:1.A proof of principle for multiplexed photoelectrochemical immunoassay of dual cardiac markers using specific enzyme tagsMultiplexed assays are capable of measuring specific biomolecules that are grouped in panels such as allergens and cardiac markers in biological matrices.It is particularly valuable in clinical laboratories if a ratio of components is required to confirm diagnostic screening or to survey the potential biochemical recurrence.In the second chapter,we proposed a simultaneous PEC immunoassay of dual cardiac markers based on specific enzyme tags.On the basis of the CdS quantum dots functionalized TiO2 nanotubes electrode,alkaline phosphatase(ALP)and acetylcholine esterase(AChE)were thus used to differentiate the signals of two cardiac targets in connection with the sandwich immunostructure and PEC responses to the corresponding in-situ generated electron donors.This strategy demonstrates a proof of principle for the successful integration of dual enzyme tags with PEC immunoassay that can potentially provide a general format for multiplexed PEC bioanalysis.2.An integration of simultaneous photoelectrochemical and visualized immunoassay of ?-human chorionic gonadotrophinIn the third chapter,ALP-induced biocatalytic precipitation reaction was first introduced into PEC bioanalysis,and a simultaneous PEC and visualized immunoassay has been established for the detection of ?-human chorionic gonadotrophin(?-HCG).Specifically,in the proposed system,ALP stimulated the oxidative hydrolyzing transformation of 5-bromo-4-chloro-3-indoyl phosphate(BCIP)to an indigo precipitation,generating an insulating layer that impeded the interfacial mass and electron transfer and thus the photocurrent production.Meanwhile,a visualized detection could be performed according to the change of electrode color intensity.With high sensitivity and specificity,this work presents the first general protocol for simultaneous PEC and visualized detection,which could be easily extended to addressing numerous other biomarkers.3.Nanochannels photoelectrochemical biosensorNanochannels have brought new opportunities for biosensor development.In the fourth chapter,we present the novel concept of a nanochannels PEC biosensor based on the integration of a unique CuxO nanopyramid-islands(NPIs)photocathode,an anodic aluminum oxide(AAO)membrane,and alkaline phosphatase(ALP)catalytic chemistry.In this protocol,the AAO nanochannel arrays acted simultaneously as a platform for protein loading,a vessel for biochemical reactions and the collecting of products,and also an aisle for the connection of the solid-liquid interface.After biocatalytic precipitation reaction initiated within the channels,the biosensor could be used for successful detection of ALP activity,with the PEC signal as an output and visualized signal as a verification.This study not only provided a novel paradigm for an unconventional nanochannels PEC biosensor,which can be used for general bioanalytical purposes,but also indicated that the new concept of nanochannel-semiconductor heterostructures is a step toward innovative biomedical applications.4.Photoelectrochemical biosensor for miRNA-141 in whole blood sample based on the integration of entropy-driven toehold-mediated DNA strand displacement reaction,magnetic separation process and energy transferHighly stable circulating microRNAs(miRNAs)are currently recognized as a novel potential biomarker for clinical cancer diagnosis in the early stage.However,limited by its low concentration,high sequence similarity as well as the numerous interferences in body fluids,detection of miRNA in whole blood with sufficient selectivity and sensitivity is still challenging.In the fifth chapter,we reported the integration of entropy-driven toehold-mediated DNA strand displacement(ETSD)reaction with magnetic beads(MB)toward the energy-transfer-based PEC detection of the prostate carcinoma(PCa)biomarker miRNA-141 in real blood sample.In this protocol,target extraction and amplification could be realized in a single test and ultrasensitive detection of miRNA-141 could be achieved in undiluted whole blood sample.This work proposed an innovative solution for sensitive biomolecular detection in complex biological milieu and exhibited great promise for future clinical cancer diagnosis.