Establishment And Application Of Lateral Flow Immunoassay For Sulfamethazine

Sulfamethazine(SMZ),a kind of broad-spectrum antibacterial veterinary drug,is widely used in animal husbandry.SMZ can interfere with the folate metabolism pathway of bacteria and then affect the normal growth and reproduction of bacteria.The unreasonable use of SMZ may cause its residues in animal-derived foods and accumulates in the human body through the food chain,thereby threatening human health.Therefore,it is important to establish the rapid and sensitive detection methods to monitor SMZ residues in animal-derived foods.In recent years,lateral flow immunoassay(LFIA)has been widely used in various fields including environmental monitoring,disease diagnosis,and food safety testing due to its advantages of simplicity,rapidity,and low cost.However,conventional LFIA using gold nanoparticles(Au NPs)as the signal label exhibits relatively poor sensitivity and mainly focuses on qualitative detection,which hinders its trace analysis of some low-concentration of targets.At present,it is important to select some novel nanomaterials with unique fluorescence characteristics or optical absorption as the signal labels to improve the detection performance of LFIA.In this study,three novel nanomaterials were selected as the labels to construct three novel types of LFIA,which were applied to the detection of SMZ residue.In the first chapter,the physical and chemical properties,hazards,mechanism of antibacterial,maximum residue limits,and current detection research progress of SMZ were systematically introduced.Then,the device,principle,signal labels,and sample addition methods of the detection platform of LFIA were summarized.In the second chapter,time-resolved fluorescent nanobeads(TRFN)was used as the signal label to develop a kind of LFIA for the highly sensitive detection of SMZ residues in egg,honey,and pork samples.Under the optimized working conditions,the TRFN-LFIA had limits of detection(LOD)of 0.016,0.049,and 0.029ng/mL and corresponding linear ranges of 0.05–1,0.05–5,and 0.05–1ng/mL in egg,honey,and pork samples,respectively.The recovery experiments showed that the average recoveries of TRFN-LFIA were 90.5%–113.9%,82.4%–112.0%,and 79.8%–93.4%with corresponding coefficients of variation of 4.1%–11.7%,7.5%–11.5%,and 4.8%–8.7%for egg,honey,and pork samples,respectively.Besides,45 actual samples were simultaneously analyzed by TRFN-LFIA and HPLC-MS/MS,and the two detection results were consistent,which proved that the TRFN-LFIA had good accuracy.In the third chapter,a competitive LFIA with aggregation induced emission fluorescent microsphere(AIEFM)as the signal label was established for the first time,which achieved the highly sensitive detection of SMZ residues in honey.Under the optimized working conditions,the LOD of the developed competitive AIEFM-LFIA in honey was 0.028ng/mL,and the corresponding linear range was 0.05-5ng/mL.Recovery experiments showed that the average recovery of AIEFM-LFIA was 75.6%–95.1%with coefficient of variation of 5.7%–12.9%in honey.Besides,15 honey samples were simultaneously analyzed by AIEFM-LFIA and HPLC–MS/MS,and the two detection results were consistent,which proved that the AIEFM-LFIA had good accuracy.In the fourth chapter,a novel multimodal signal readout LFIA based on dual fluorescence quenching of polydopamine nanospheres(PDANs)was developed for the highly sensitive detection of SMZ residues in chicken.Compared with traditional quenchers such as Au NPs with single fluorescence quenching ability,PDANs have dual fluorescence quenching ability,which could simultaneously quench the excitation light and emission light of three fluorescence donors(AIEFM;fluorescent microsphere,FM;and quantum dot bead,QB).The two quenchers were simultaneously applied to the fluorescence quenching LFIA,and the results showed that the change values of fluorescence intensity based on PDANs-LFIA(?F_AIEFM=2315,?F_FM=979,?F_QB=910)were all greater than those based on Au NPs-LFIA(?F_AIEFM=1722,?F_FM=833,?F_QB=520).These results proved that PDANs can be used to improve the quenching efficiency and sensitivity of fluorescence quenching LFIA.Under the optimized working conditions,the LOD and visual LOD of LFIA based on PDANs quenching AIEFM for the detection of SMZ in chicken were 0.043 and 0.5ng/mL,respectively.In the fifth chapter,three kinds of LFIA developed in this paper for detecting SMZ were systematically summarized,and the possible methods for improving the detection performance of LFIA in the future were proposed.