Preparation Of Immunochromatographic Test Strips Based On Different Probes And Detection Mode

With the development of economic level,food safety issues have received more and more attention from the people and the government,and become a hot national topic.Under this situation,some of food safety testing technologies have also obtained a certain degree of development.Immunochromatographic assay(ICA)is based on the combination of tomography technology and the antigen-antibody specific interaction.The typical characteristics of ICA are simplicity,quickness,no requirements for professional personnel and equipment,low cost,convenience,etc.In 2015,the newly revised Food safety law of the People's Republic of China has clarified the important status of rapid detection method for food safety monitoring.Traditional ICA using colloidal gold(CG)as the signal output label displayed low sensitivity,generally can only be used for qualitative analysis.As the miniaturization of optical testing instrument and the emergence of new signal labels,more and more new fluorescent labeling materials were reported.However,few of the ICA products based on new fluorescent labels were applied into practical application and the pros and cons of these fluorescent labels were not entirely understood.Therefore,in this study,various ICAs based on different labels for detection of Aflatoxin B1(AFB1)were used to clarify and evaluate the properties of labels and their corresponding industrialization prospect.Meanwhile,since currently there were only qualitative or quantitative test can be performed on ICA strips,clenbuterol hydrochloride(CLE)was selected as the model analysis target to explore and develop qualitative and quantitative ICA strips.The first chapter introduced the physical and chemical properties,hazards,pollution,detection methods of AFB1.The application of different labels in ICA,the development background,and significance of simultaneously qualitative and quantitative strip were summarized.The chapter 2 systematically analyzed that labels such as the CG,fluorescent microspheres(FM),time-resolved fluorescence microspheres(TRFN),and quantum dots microspheres(QM)were adopted to prepare ICAs based on competitive model for detection of AFB1.Furthermore,the optimum parameters of the ICAs based on four kinds of labels namely,CG-ICA,FM-ICA,TRFN-ICA,and QM-ICA were optimized in this chapter.The half maximal inhibitory concentrations(IC50)of these ICAs for detection of AFB1 in core samples were 0.15 ng/mL(CG-ICA),0.08 ng/mL(FM-ICA),0.014 ng/mL(TRFN-ICA)and 0.03 ng/mL(QM-ICA),respectively.The sensitivity of TRFN-ICA for detection of AFB1 was the highest one,however,the QM-ICA displayed the best precision and stability among four ICAs.we believed QM-ICA was suitable for industrial production,which has the advantages of high sensitivity,easy to label and separation,and good stability.The detection results indicated that the IC50 of QM-ICA for detection of AFB1 in different environments were from 0.03?0.05 ng/mL with a wide detection linear range of 0.01-0.5 ng/mL.The detection time was 15 min.QM-ICA has a good application value and market prospect for detection of small molecules.In the chapter 3,CG and TRFN probes were used to establish an integrated immunochromatographic assay to qualitatively and quantitatively detection CLE.The best experimental conditions for the two probes in separate ICAs were obtained by optimizing the antibody labeling concentration,the amount of antigen,and the concentration of probe.When the CG and TRFN probes co-existed in the ICA,the latter had no effect on the sensitivity of qualitative detection of the CG probe-based ICA.However,the CG probe optimized the linear range of quantitative detection in the TRFN probe-based ICA.The integrated test strip can be used for qualitative and quantitative detection of CLE in one step.When the amount of antigen reached 0.4 mg/mL,the CG probe concentration reached 1.2 ?g/mL,and the TRFN probe concentration reached 0.68?g/mL.The qualitative sensitivity of the integrated ICA was 0.5 ng/mL and its quantitative limit of detection was 0.04 ng/mL with a detection range of 0.1-2.7 ng/mL.This developed method is of great significance for large-scale samples screening and positive monitoring in the field of food safety testing.The fourth chapter summarized the full study.This study systematically and comprehensively analyzed the four probes in industrial applications and exploited the integrated use of dual-signal probes.It is of great significance to evaluate the industrialization prospects of different probes.The integration of labels broadened the functionality and application range of rapid detection products,which has a guiding role in product development.