Fluorescence Quenching: Theory and Applications
Fluorescence Quenching: Theory and Applications
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DOI：
10.1007/0-306-47058-6_2
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发表时间：
2002
期刊：
影响因子：
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通讯作者：
M. Eftink
中科院分区：
文献类型：
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作者：
M. Eftink
溶质荧光猝灭反应在20世纪60年代末和70年代初首次应用于生物化学问题，（1-7）从那时起，它们就成为研究蛋白质，膜和其他大分子组装体的非常有价值的研究工具。猝灭反应易于进行，只需要少量样品，通常是非破坏性的，并且可以应用于几乎任何具有内在或外在荧光探针的系统。然而，最重要的特征是这些反应可以提供的信息的价值。使用猝灭剂如分子氧、丙烯酰胺或碘离子的溶质猝灭反应提供了关于荧光基团在大分子结构中的位置的信息。位于较大结构的表面上的荧光团将相对容易接近溶解在水相中的溶质猝灭剂。从结构的表面去除的荧光团将被猝灭剂猝灭至较低程度。因此，猝灭反应可用于探测大分子组装体的形貌特征，并检测可能由变化的条件或添加试剂引起的任何结构变化。此外，在某些情况下，猝灭反应可以提供有关构象波动的信息。在2.3节和2.4节中，我将讨论在蛋白质、膜和核酸研究中使用溶质猝灭剂的几个例子。溶质荧光猝灭反应也可用于选择性地改变样品的荧光性质，以便解析贡献或帮助测量数据。为了详细说明这一点，考虑荧光的不同特性：量子产率、激发和
Solute fluorescence quenching reactions were first applied to biochemical problems in the late 1960s and early 1970s,(1–7) and since that time they have been a very valuable research tool for studies with proteins, membranes, and other macromolecular assemblies. Quenching reactions are easy to perform, require only a small sample, usually are nondestructive, and can be applied to almost any system that has an intrinsic or extrinsic fluorescence probe. The most important characteristic, however, is the value of the information that these reactions can provide. Solute quenching reactions, using quenchers such as molecular oxygen, acrylamide, or iodide ion, provide information about the location of fluorescent groups in a macromolecular structure. A fluorophore that is located on the surface of a larger structure will be relatively accessible to a solute quencher that is dissolved in the aqueous phase. A fluorophore that is removed from the surface of a structure will be quenched to a lesser degree by the quencher. Thus, the quenching reaction can be used to probe topographical features of a macromolecular assembly and to sense any structural changes that may be caused by varying conditions or the addition of reagents. In addition, quenching reactions can, in some situations, provide information about conformational fluctuations. In Sections 2.3 and 2.4 I will discuss several examples of the use of solute quenchers in studies with proteins, membranes, and nucleic acids. Solute fluorescence quenching reactions can also be used to selectively alter the fluorescence properties of a sample in order to resolve contributions or aid in the measurement of data. To elaborate on this point, consider the different characteristics of fluorescence: the quantum yield, excitation and