Ozinskas A J, Malak H, Joshi J, Szmacinski H, Britz J, Thompson R B, Koen P A, Lakowicz J R
Becton Dickinson Advanced Diagnostics, Baltimore, Maryland 21209.
Anal Biochem. 1993 Sep;213(2):264-70. doi: 10.1006/abio.1993.1419.
We describe a homogeneous competitive model immunoassay for determination of thyroxine by multifrequency phase-modulation fluorescence. Using a nonradiative energy transfer transduction mechanism, B-phycoerythrin conjugated to thyroxine is the energy donor and a carboxymethylindocyanine dye conjugated to anti-thyroxine antibody is the energy acceptor. Energy transfer from B-phycoerythrin to the acceptor results in a decreased lifetime and/or phase angle. The fluorescence lifetime change reflects the extent of energy transfer. In the competitive immunoassay format, the donor-thyroxine conjugate and an analytical sample of thyroxine compete for acceptor-antibody binding sites, resulting in a phase angle change which is dependent on the amount of thyroxine in the sample. Dose-response curves of phase angle versus thyroxine concentration are comparable to steady-state intensity curves. Since phase-modulation lifetime measurements are largely independent of total signal intensity, sources of optical interference are minimized. The potential for whole blood measurements exists since the energy transfer lifetime method can be extended to longer wavelengths.
我们描述了一种用于通过多频相位调制荧光测定甲状腺素的均相竞争模型免疫分析方法。利用非辐射能量转移转导机制，与甲状腺素偶联的B-藻红蛋白作为能量供体，与抗甲状腺素抗体偶联的羧甲基吲哚菁染料作为能量受体。从B-藻红蛋白到受体的能量转移导致寿命缩短和/或相角减小。荧光寿命变化反映了能量转移的程度。在竞争免疫分析形式中，供体-甲状腺素偶联物和甲状腺素分析样品竞争受体-抗体结合位点，导致相角变化，该变化取决于样品中甲状腺素的量。相角与甲状腺素浓度的剂量-反应曲线与稳态强度曲线相当。由于相位调制寿命测量在很大程度上与总信号强度无关，因此光学干扰源被最小化。由于能量转移寿命方法可以扩展到更长波长，因此存在进行全血测量的潜力。
