使用4,7-双（氯磺苯基）-1,10-菲咯啉-2,9-二羧酸（BCPDA）的铕螯合物进行时间分辨荧光。标记程序及其在免疫测定中的应用。
Time-resolved fluorescence using a europium chelate of 4,7-bis-(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA). Labeling procedures and applications in immunoassays.
作者信息
Diamandis E P, Morton R C
机构信息
Cyber Fluor Inc., Toronto, Ontario, Canada.
出版信息
J Immunol Methods. 1988 Aug 9;112(1):43-52. doi: 10.1016/0022-1759(88)90031-2.
We describe optimal conditions for protein labeling with a new fluorescent probe, 4,7-bis-(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA). The labeled proteins are suitable for time-resolved fluorometric applications because BCPDA forms fluorescent complexes with Eu3+ which exhibit very long fluorescence lifetimes. Labeling parameters such as the organic solvent used, pH, protein concentration, BCPDA excess and incubation times, were optimized accordingly. Excess BCPDA was removed by gel filtration and labeled proteins were characterized by absorbance and fluorescence measurements. The effect of labeling on the biological (binding) activity of the proteins streptavidin, avidin, monoclonal and polyclonal antibodies was also studied. It is shown that the labeled antibodies can be used for time-resolved fluoroimmunoassay applications.
我们描述了使用新型荧光探针4,7-双（氯磺苯基）-1,10-菲咯啉-2,9-二羧酸（BCPDA）进行蛋白质标记的最佳条件。标记后的蛋白质适用于时间分辨荧光分析应用，因为BCPDA与Eu3+形成具有很长荧光寿命的荧光复合物。相应地优化了标记参数，如所用有机溶剂、pH值、蛋白质浓度、BCPDA过量情况和孵育时间。通过凝胶过滤去除过量的BCPDA，并通过吸光度和荧光测量对标记后的蛋白质进行表征。还研究了标记对链霉亲和素、抗生物素蛋白、单克隆和多克隆抗体等蛋白质生物（结合）活性的影响。结果表明，标记后的抗体可用于时间分辨荧光免疫分析应用。
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