| Point of Care Testing(POCT)is a type of medical diagnostic test performed in a place where patients outside the hospital or outside the laboratory are diagnosed,usually by non-laboratory professionals,who can help people get immediate health care.Information to provide a reference for clinical diagnosis for the next decision.Instant detection has the advantages of expanding diagnostic accessibility,small sample size,low cost and fast analysis.In addition,with the development of smart phones and communication industry,real-time detection combined with Internet+mode can realize convenient and fast remote operation.Medical care,which has important positive significance for solving imbalances in the distribution of medical resources in China and the world.At present,the most widely used instant detection equipment is blood glucose meter.Most blood glucose meters on the market are based on electrochemical principle.In addition to the necessary test strips,additional electrochemical detection devices are needed,which are expensive.Therefore,this dissertation aims to develop a novel electrogenerated chemiluminescence(ECL)bioanalytical system for enzyme-catalyzed reactions on paper substrates using smart phones as a detection platform for the immediate detection and screening of disease markers.The results of this project will promote the development of real-time detection devices and methodological research,and provide new ideas and new methods for people’s health inspection in economically underdeveloped areas in accordance with the times,and have important scientific and social significance.This study was funded by the project of national natural science foundation of China,"study on some basic problems of electrochemiluminescence biosensor"(21475082,January 2015--December 2018)and"study on field detection methods of electrochemiluminescence biosensor"(21775098,January 2018--December 2021).This paper consists of two parts:introduction and research report.The first part is the introduction,briefly introduces the basic principles of electrochemiluminescence analysis methods and commonly used electrochemiluminescence systems,glucose electrochemical enzyme biosensors and peptide biosensors,research progress of paper-based real-time detection and the application of smart phones for instant detection.The research progress,and finally put forward the research background,research ideas,research purposes and research content of this thesis.The second part is the research report part,and the research report part consists of the following two parts:First,an electrogenerated chemiluminescence biosensing method for the detection of blood glucose incorporating a paper-based microfluidic sensor and a smartphone is developed.Glucose reacts specifically with glucose oxidase to produce hydrogen peroxide.Hydrogen peroxide acts as a co-reactant to produce an electrogenerated chemiluminescence reaction with luminol.The concentration of glucose in the sample solution is determined by measuring the intensity of electrogenerated chemiluminescence.Based on this principle,this paper developed a paper-based microfluidic chip for detect blood glucose by used smartphone,and established an electrogenerated chemiluminescence biosensing method for detecting blood glucose.Preparation of paper-based microfluidic electrogenerated chemiluminescence sensor devices on selected chromatographic papers(Waterman 1Chr),including sample pads(containing glucose oxidase GOD),electrogenerated chemiluminescence reaction pads,wax hydrophobic bands and fixed ECL reagents Two electrodes(the pencil draws the working electrode and the counter/reference electrode for printing the conductive silver paste,and the ECL reagent luminol is applied to the surface of the working electrode).When a blood sample(about 15 μL)is dropped on the fixed part of the sample pad,the red blood cells and proteins in the sample are adsorbed on the sample pad,and hydrogen peroxide and other small molecules generated by the reaction of glucose and enzyme further flow forward to the paper based on capillary action.On the base electrode.The potential of the working electrode was controlled to be+1.5 V(vs.Ag),and the resulting electrogenerated chemiluminescence was taken with a camera on a smartphone(iPhone6S Plus).The RGB value is read by the software in the smartphone for the electrochemiluminescence image taken,and the read RGB value is applied to quantify the glucose concentration in the sample.The experimental conditions of the prepared glucose electrogenerated chemiluminescence paper-based microfluidic chip were selected,and the paper-based materials,electrode materials,glucose oxidase and luminol fixation methods were selected.The optimal luminol concentration and optimum rumm were selected.The conditions of luminol’s pH,optimum glucose oxidase concentration and optimum electrogenerated chemiluminescence potential of the potentiostatic method were optimized.The results obtained by this method were compared with the results of the mainstream blood glucose meter on the market.The experimental results show that the RGB value is proportional to the logarithm of the glucose concentration in the sample,and the linear range is 2.4～30.9 mM,which meets the requirements of clinical testing.The results of actual blood glucose concentration measured by this method are consistent with those of mainstream blood glucose meters in the market.Second,the exploratory study of bio-cleavage ECL paper-based microfluidic sensor.Based on the above work,we hope that the method of detecting the ECL of paper chips by smart phones should be promoted to the lower content of the target by detecting substances with higher content in samples such as glucose.The prostate specific antigen(PSA)was used as the target analyte,and the specific sequence polypeptide(GSGSGSHSSKLQK)was used as the molecular recognition substance,and the pyridine derivative Ru(bpy)2(mcbpy-O-Su-ester)(PF6)2 was used as the molecular recognition substance.The ECL signal substance has been exploratoryly researched on the design and fabrication of bio-cleavage ECL paper chips.The bio-cleavage ECL paper-based microfluidic chip is composed of a sample pad,a reaction zone,a separation zone,a paper-based electrode and a support pad.When the target PSA reacts with MBs-SA-peptide-Ru1,the PSA cleaves the specific site of the polypeptide(between QK),and the cut Rul-K separates from the magnetic beads and migrates to the surface of the paper-based electrode with the mobile phase.The ECL reaction was carried out in the presence of the co-reactant TPrA to measure the target content.Experimental results show that the design is feasible.Signal magnetic beads probe was synthesized and its characterization,biological functional magnetic beads paper cutting chip of design parts material and the mobile phase selection,examines the electrogenerated chemiluminescence response mat the relationship between the electrochemical luminescence intensity and concentration of Rul,examines the magnetic particle size and peptide sequence of electrogenerated chemiluminescence response cushions to the influence of the electrogenerated chemiluminescence intensity.The lower content results in less ECL,and the paper blockage(only about 10%of the light is transmitted through the paper)causes the detected signal changes to be less sensitive,and we are still exploring the use of new paper-based materials,Electrode materials,etc.to solve the above problems. |
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