Lakowicz J R, Keating S
J Biol Chem. 1983 May 10;258(9):5519-24.
Phase-sensitive detection of fluorescence was used to measure the fractional binding of an amphipathic derivative of indole to detergent micelles. By selection of the detector phase angle, the emissions from the free and bound indole could be individually suppressed. The phase-sensitive intensity was then proportional to the concentrations of the bound and free indole, respectively. Also, the ratio of the phase-sensitive intensities, recorded at the phase angles for suppression, revealed the ratio of the fluorophore present in each environment. As the detergent concentration was increased, the measurements revealed a cooperative formation of detergent micelles and binding of the indole derivative to these micelles. The theory and procedures for this specific analysis can be applied to the analysis of any association reaction in which one of the species is fluorescent, and in which the fluorescence lifetime is altered by the binding process. An advantage of this procedure is that binding can be quantified without physical separation of the individual species.
利用荧光相敏检测来测量吲哚的两亲衍生物与去污剂胶束的分数结合。通过选择检测器相角，可以分别抑制游离吲哚和结合吲哚的发射。然后，相敏强度分别与结合吲哚和游离吲哚的浓度成正比。此外，在抑制相角下记录的相敏强度之比揭示了每种环境中存在的荧光团的比例。随着去污剂浓度的增加，测量结果显示去污剂胶束的协同形成以及吲哚衍生物与这些胶束的结合。这种特定分析的理论和程序可应用于任何一种物种具有荧光且荧光寿命因结合过程而改变的缔合反应的分析。该程序的一个优点是无需对各个物种进行物理分离即可对结合进行定量。
