Wang Guilan, Yuan Jingli, Hai Xiaodan, Matsumoto Kazuko
Department of Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China.
Talanta. 2006 Aug 15;70(1):133-8. doi: 10.1016/j.talanta.2005.11.052. Epub 2006 Jan 6.
A sensitive homogeneous time-resolved fluoroimmunoassay method for 3,5,3'-triiodo-l-thyronine (T3) based on the fluorescence resonance energy transfer (FRET) from a fluorescent Eu(3+) complex, {[(4,6-dichloro-1,3,5-triazin-2-yl)amino-biphenyl-4'''-yl]-2,2':6',2''-terpyridine-6,6''-diyl}bis(methylenenitrilo) tetrakis(acetate)-Eu(3+) (DTBTA-Eu(3+)) (lambda(ex,max)=335nm, lambda(em,max)=615nm), to an organic fluorescence dye Cy5 has been developed. The new assay system combined the use of DTBTA-Eu(3+)-labeled T3-bovine serum albumin (BSA) conjugate and Cy5-labeled anti-T3 monoclonal antibody for a competitive-type immunoassay. After the competitive reactions of DTBTA-Eu(3+)-labeled T3-BSA and T3 sample with Cy5-labeled anti-T3 antibody, the T3 concentration was measured with a time-resolved mode by monitoring the sensitized emission of Cy5 derived from FRET in a homogeneous format. The method gives the detection limit of 0.26ng/ml. The coefficient variations of the method are less than 2.0% and the recoveries are in the range of 80-111% for serum sample measurement. The concentrations of T3 in 30 human serum samples were determined, and the results were compared with those of the independently determined by a radio-immunoassay method. A good correlation was obtained with a correlation coefficient of 0.989.
基于荧光共振能量转移（FRET），从荧光铕（Eu(3+)）配合物{[(4,6-二氯-1,3,5-三嗪-2-基)氨基-联苯-4'''-基]-2,2':6',2''-三联吡啶-6,6''-二基}双(亚甲基腈基)四乙酸铕（Eu(3+)）（DTBTA-Eu(3+)）（激发波长最大值λ(ex,max)=335nm，发射波长最大值λ(em,max)=615nm）到有机荧光染料Cy5，开发了一种用于检测3,5,3'-三碘-L-甲状腺原氨酸（T3）的灵敏的均相时间分辨荧光免疫分析方法。新的分析系统将DTBTA-Eu(3+)标记的T3-牛血清白蛋白（BSA）偶联物和Cy5标记的抗T3单克隆抗体用于竞争型免疫分析。在DTBTA-Eu(3+)标记的T3-BSA和T3样品与Cy5标记的抗T3抗体进行竞争反应后，通过在均相形式下监测FRET产生的Cy5的敏化发射，以时间分辨模式测量T3浓度。该方法的检测限为0.26ng/ml。对于血清样品测量，该方法的变异系数小于2.0%，回收率在80-111%范围内。测定了30份人血清样品中T3的浓度，并将结果与用放射免疫分析方法独立测定的结果进行比较。获得了良好的相关性，相关系数为0.989。
