Preparation And Immunochromatographic Application Of Long Life Fluorescent Rare Earth Doped Nanocrystals

Time-resolved fluoroimmunoassay is an immunoassay technology developed based on the unique fluorescent properties of rare earth elements.It combines the advantages of radioimmunoassay technology,enzyme-linked immunoassay technology and ordinary fluorescence immunoassay technology.The high sensitivity,strong specificity,good stability,wide linear range,long fluorescence lifetime,simple operation and non-radioactive characteristics of time-resolved fluoroimmunoassay shows a good application prospect in the field of immunoassay.In this paper,a rare earth-doped nanocrystal LaF₃:Eu³⁺with high crystallinity,good monodispersity,and long fluorescence lifetime is prepared via hydrothermal method.The surface of LaF₃:Eu³⁺nanocrystal is functionally modified by polyacrylic acid(PAA),and gadolinium ion(Gd³⁺)and(ZMA)are used as sensitizers for luminescent center europium ion(Eu³⁺)to improve the luminous efficiency of LaF₃:Eu³⁺nanocrystals.At the asme time,a time-resolved fluorescence immunoassay technology based on rare earth fluorescent microspheres is established for the detection of new coronavirus neutralizing antibodies in serum samples.The main research contents and conclusions of this paper as follow:(1)LaF₃:Eu³⁺nanocrystals with different europium ion(Eu³⁺)doping degrees are prepared by hydrothermal method.Their crystal structure,morphology,and size are basically the same,and they all present a relatively regular hexagon-like structure,the particle sizes are about 50 nm.The fluorescence spectrum of LaF₃:Eu³⁺nanocrystals show that their excitation and emission wavelengths are consistent with the luminescent center Eu³⁺ions,and the fluorescence lifetime are as high as 3 to 4 ms.Besides,LaF₃:Eu³⁺@PAA nanocrystals have successfully been functionalized with polyacrylic acid(PAA),which will lay the foundation for their subsequent application research.(2)The inorganic sensitizer gadolinium ion(Gd³⁺)and activator europium ion(Eu³⁺)are co-doped in the matrix of lanthanum fluoride nanocrystals.The effect of Gd³⁺ion doping on the structure and fluorescence performance of nanocrystals are studied.It is found that the the fluorescence lifetime of nanocrystals are prolonged with the doping of Gd³⁺ions,from 4.07 milliseconds(LaF₃:0%Gd³⁺,Eu³⁺)to 5.92 milliseconds(LaF₃:40%Gd³⁺,Eu³⁺).However,the sensitization effect of Gd³⁺ion is not good,and the stability of the nanocrystals is decreased.The organic sensitizer ZMA and non-aqueous auxiliary ligand FZB are modified to the surface of the LaF₃:Eu³⁺nanocrystals.By replacing the fluorescence quenching group-OH with FZB,the fluorescence performance of nanocrystals is effectively enhanced by the resonance energy transfer between ZMA and Eu³⁺.But the modification of ZMA and FZB significantly shortens the fluorescence lifetime of nanocrystals.(3)A time-resolved fluorescence immunochromatography method for the detection of COVID-19 neutralizing antibody is established.The IC₅₀of the blocking method and the competition method based on rare earth fluorescent microspheres is 0.82?g/m L and4.13?g/m L,respectively.The immunoassay conditions of serum samples are optimized,and the best detection conditions are obtained:the dilution ratio of serum is 1:25,the p H is7.0,the content of BSA is 2%,and the content of Tween-20 is 0.6%.