A Novel Bifunctional Europium Chelate Applied In The Research Of Time-Resolved Fluoroimmunoassay System

In recent years there has been increased interest in the development of new, simple, sensitive, and specific immunoassays for the quantification and detection of analytes of clinical and biological importance. Among all the labeled immunoassays, TRFIA methodology has developed so rapidly and has become a new milestone in the development of labeled immunoassays and clinical medicine. TRFIA holds great promise for immunoassays, owing to its unique advantages, such as high sensitivity, easy automation, less susceptibility to matrix interference, and simultaneous multilabeling. TRFIA takes full advantage of the unique fluorescent characteristics of trivalent rare-earth ions and their chelates, such as long lifetime and a large Stokes’ shift. These advantages contributed to the lower background and improved detection sensitivity when the immunoassay was quantified using TRFIA.In this study, a novel bifunctional chelate Eu(TTA)3(5-NH2-phen)(ETNP) with a long fluorescence lifetime and intense luminescence was used to label the goat anti-human IgG as a biomarker, and a new TRFIA protocol for quantification of human IgG has been established by using sandwich-type immureaction on the glass slides. The chelate ETNP could be tight coupled with the IgG and the fluorescence intensity could be directly measured on a solid support after several washing steps. The fluorescent signal obtained by fluorescence detection instrument is proportional to the concentration of human IgG, which can be used as quantitative analysis basis by this method. A minimum of5μg/L could be detected which was greatly improved in sensitivity, comparing with RIA of1000μg/L and ELISA of200μg/L in human IgG detection. To the best of our knowledge, this was the first time that a sensitive and specific immunofluorometric method with antigen detection, was established by using the novel europium chelate ETNP as the fluorescence label. This method has so many advantages such as simple operation, low detection limitation, high sensitivity. Based on the detection system of human IgG, the author also uses the bifunctional chelates ETNP to develop alpha-fetoprotein (AFP) quantitative detection system, which has achieved good results. The lowest concentration of AFP could be detected is22μg/L, and with good repeatability.The establishment of the new protocol using ETNP as the label material has greatly improved the quantitative detection of human IgG. The authors also contend that the method has enormous promise for further applications to antigen and antibody detection.