Dual-color fluorescence cross-correlation spectroscopy for multicomponent diffusional analysis in solution
Dual-color fluorescence cross-correlation spectroscopy for multicomponent diffusional analysis in solution
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DOI：
10.1016/s0006-3495(97)78833-7
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发表时间：
1997-04-01
影响因子：
3.4
通讯作者：
Rigler, R
中科院分区：
文献类型：
--
作者：
Schwille, P;MeyerAlmes, FJ;Rigler, R
本文描述了一种新的实验方案，通过荧光相关分析来跟踪纳摩尔浓度范围内荧光标记分子的扩散和化学反应体系。在此提供的双色荧光互相关光谱中，可以并行跟踪溶液中两种荧光物质及其反应产物的浓度和扩散特性。与普通的单色荧光自相关系统相比，通过使用两种不同标记的反应配偶体，研究反应产物的时间演化的选择性显着增加。在这里，我们开发了在共焦双光束荧光相关光谱装置中进行测量的理论和实验基础，并讨论了有利于互相关分析的条件。解释了使用两条不同标记的互补链进行 DNA-DNA 复性动力学的测量原理。反应产物的浓度可以直接从互相关幅度确定。
The present paper describes a new experimental scheme for following diffusion and chemical reaction systems of fluorescently labeled molecules in the nanomolar concentration range by fluorescence correlation analysis. In the dual-color fluorescence cross-correlation spectroscopy provided here, the concentration and diffusion characteristics of two fluorescent species in solution as well as their reaction product can be followed in parallel. By using two differently labeled reaction partners, the selectivity to investigate the temporal evolution of reaction product is significantly increased compared to ordinary one-color fluorescence autocorrelation systems. Here we develop the theoretical and experimental basis for carrying out measurements in a confocal dual-beam fluorescence correlation spectroscopy setup and discuss conditions that are favorable for cross-correlation analysis. The measurement principle is explained for carrying out DNA-DNA renaturation kinetics with two differently labeled complementary strands. The concentration of the reaction product can be directly determined from the cross-correlation amplitude.