Development Of Fluorescence Immunoassay For Two Emerging Contaminants In The Environments

In recent years,due to Emerging Contaminants(Ecs)in the ecological environment have posed a potential or major threat to human health.So,it has received extensive attention in the field of environmental chemistry.Sulfonamide(SAs)and Tetrabromobisphenol A(TBBPA)are two typical examples of ECs.Although the two types of substances in the environment are present in lower concentrations,they can still enter the environment and organisms in various ways,which have a negative impact on the ecological environment and human health.Therefore,it is necessary to establish a sensitive,fast and efficient analysis method for investigating these pollutes.Fluorescence immunoassay based on fluorescence technology and antigen-antibody specific recognition has a series of advantages:simple operation process,no complicated sample preparation,high throughput,etc,which can meet the rapid and sensitive analysis of trace pollutants in a short time.Based on silver nanoclusters(AgNCs),FAM-DNA-functionalized CdSe/ZnS QDs,CdTe/CdS QDs,three fluorescence immunoassay methods were established.The details are as follows:(1)Based on the quenching of I-to silver clusters(AgNCs),a novel indirect competitive fluorescence immunoassay was developed.This work regards SAs in environmental samples as detecting target,first,the alkaline phosphatase(ALP)was labeled on the secondary antibody(Ab2).And after the competition step,the magnesium ascorbyl phosphate could be catalyzed to produce the ascorbic acid under the catalysis of ALP.Subsequently,the I2 was introduced and further reduced to I-in the presence of ascorbic acid,triggering the fluorescence quenching of AgNCs dispersed in isopropanol(IPA)buffer.More importantly,trace I-could lead to an obvious reduction in fluorescence signal,indicating the sensitivity of this method would be greatly improved.Under the optimal condition,the improved method for the SAs detection has a lower detection of limit(LOD,0.05 μg L-1)and a wider range(0.1471.71 μg L-1).After the evaluation,the fluorescence ELISA proposed in this work has satisfactory accuracy and reliability(recoveries,84.18-118.6%;CV,2.03%to 7.64%),illustrating good performance and great potential for the detection of trace pollutant in the environment.(2)In the previous work,although the fluorescence immunoassay method is sensitive,However,in the analysis process,fluorescent probes based on a single wavelength are more susceptible to local environment,concentrations of probe and excitation source fluctuations,so their application was limited.Therefore,a more stable method is needed.In this work,a simple indirectly competitive ratiometric fluorescent immunoassay was designed based on FAM-DNA-functionalized CdSe/ZnS QDs.This work regards TBBPA in environmental and food samples as detecting target,at the detection system,the catalase(CAT)was lablled on the secondary antibody(Ab2),which serves as a "controller" of the H2O2 concentration.After the competitive binding step,the emitted red fluorescence(excitation at 490 nm)from FAM-DNAfunctionalized CdSe/ZnS QDs could be effectively quenched by the H2O2 added.Under the optimized conditions,the limit of detection(LOD)reached 0.118 μg L-1 with a linear range of 0.34-45.34 μg L-1,which was approximately one order of magnitude lower than that by HRP-based traditional ELISA.Furtheremore,the combination of the dual-output ratiometric fluorescence assays with ELISA improved the inherent built-in rectification to the environment,brought about the satisfactory accuracy and precision(recoveries,83.16-112.4%;CV,2.42-7.28%),indicating great potential for the determination of emerging contaminants from food and environmental samples.(3)Based on review and summary of the previous work,although the fluorescent ELISA analysis(FELISA)method has better sensitivity and higher accuracy than the traditional ELISA,it still has certain limitations in real-time detection.Therefore,it is necessary to develop a more convenient and sensitive method.In this study,Based on CdTe/CdS QDs,a fluorescence immunochromatographic test strip assay(FITSA)was developed for sensitive detection of SAs and TBBPA.Under the optimized conditions,the limit of detection(LOD)reached 1.47 μg L-1 and 2.55 μg L-1,respectively,The whole analysis process of this method could be completed in 20 min without using other complex instruments.At the same time,it also provides some experience for the development of rapid,low-cost and highly sensitive analytical methods.