Construction And Application Of Lateral Flow Immunochromatographic Strip And Fluorescence Sensor For Estradiol

Estradiol is a steroid estrogen,which has a strong endocrine disrupting effect.It can enter the body through the enrichment of the food chain,causing abnormalities in the reproductive system and immune system,and even leading to organ cancer.Studies have shown that there are varying degrees of estradiol pollution in dairy products,meat products,and environmental water samples near farms,underground and surface.Therefore,it is of great significance to develop a simple,rapid and sensitive detection method for estradiol to ensure food safety and human health.Fast inspection technology can realize on-site detection and a large number of samples screening with a fast detection speed and no need of complex instruments and professional personnel,so it has a broad market demand.As a fast detection technology,lateral flow immunochromatography strip has the advantages of simple operation,high efficiency and low cost.But the traditional colorimetric strip has low sensitivity and poor stability,it is difficult to meet the requirements of trace detection of estradiol.In this paper,fluorescence and temperature were used as signals to establish two kinds of lateral flow immunochromatographic strips to realize rapid and sensitive detection of estradiol.And the application of aptamer as recognition factor was explored in estradiol detection.The main research contents and results of this paper are as follows:(1)In order to solve the problems of low sensitivity and poor stability of colorimetric strip,a competitive fluorescence lateral flow immunochromatography method was developed for the rapid detection of estradiol.Based on the specific recognition of antigens and antibodies,a competitive fluorescence lateral flow immunochromatographic strip was constructed using europium-doped fluorescent microspheres labeled with antibodies as the signal probe.Under the optimized conditions,the T-zone intensity showed a good linear relationship with estradiol concentrations in the range of 0.1-25 ng/m L(R~2=0.9949).The detection limit was 0.1 ng/m L.Compared with the traditional colorimetric strip,this method can realize the quantitative analysis of estradiol with higher sensitivity.The recoveries of water and milk samples were86.1%?116.4%and 99.0%-112.8%,respectively.(2)In order to avoid the background fluorescence interference of fluorescent strip and further improve the sensitivity,a thermal analysis immunochromatography strip was established to realize the visualization and thermal analysis of estradiol.Au-black phosphorus nanosheet composites(Au-BPNs)were synthesized at room temperature and labeled with antibodies as a signal probe.A thermal analysis lateral flow immunochromatographic strip was constructed based on the photothermal properties of Au-BPNs.Under the optimized conditions,the thermal analysis method showed a good linear relationship in the range of 0.05-50 ng/m L(R~2=0.9979).The detection limit of thermal analysis was 0.05 ng/m L,and the visual detection limit was 5 ng/m L.Compared with fluorescent strip,the detection limit of this method is reduced and the linear range is increased.The recoveries of water samples,milk powder and milk samples were respectively 89.0%-116.4%,89.3%-95.8%and 101.1%-125.2%.(3)In order to avoid the use of antibodies and develop a non-immunological rapid detection method for E2,a fluorescence aptamer sensor was established to detect estradiol.The tertiary structures,binding sites,binding forces and binding specificities of 22,35 and 75-mer aptamers to estradiol were investigated by molecular simulation technology.After that,the 22-mer estradiol aptamer labeled with FAM was selected as fluorescence donors.With BPNs as fluorescence receptors,a fluorescence aptamer sensor was constructed based on fluorescence resonance energy transfer for the detection of estradiol.Under the optimized conditions,this method could achieve the quantitative detection of estradiol in the range of 3.3-60 ng/m L.The detection limit was 1.0 ng/m L.And it can achieve specificity detection.The recoveries of tap water and milk samples were 91.2%-104.8%and 87.5%-104.3%,respectively.It lays a theoretical foundation for the further development of aptamer-based lateral flow strip.