Chemical calcium indicators.
Chemical calcium indicators.
复制标题
化学钙指标。
DOI：
10.1016/j.ymeth.2008.09.025
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发表时间：
2008-11
期刊：
影响因子：
4.8
通讯作者：
Lechleiter, James D.
中科院分区：
文献类型：
--
作者：
Paredes, R. Madelaine;Etzler, Julie C.;Watts, Lora Talley;Zheng, Wei;Lechleiter, James D.
我们对Ca 2+信号传导的潜在机制的理解以及我们对其在细胞过程中无处不在的作用的理解，在很大程度上是由于荧光Ca 2+指标的发展而迅速发展的。在本章中，我们讨论了一些最常见的化学Ca 2+指标，被广泛用于细胞内Ca 2+信号的调查。每种染料的优点、局限性和相关程序将被介绍，包括它们的光谱质量、离解常数、化学形式、装载方法和设备，以实现最佳成像。现在可用的化学指示剂允许在非常大的范围（50 μM）内检测细胞内Ca 2+<50 nM to >。较高亲和力的指示剂可用于定量胞质溶胶中的Ca 2+水平，而较低亲和力的指示剂可被优化用于测量具有较高浓度的亚细胞区室中的Ca 2+。指示剂可分为单波长或比率染料。这两类都需要特定的激光器、滤光片和/或检测方法，这些方法取决于它们的光谱特性，并且这两类都有优点和局限性。单波长指示剂通常非常明亮，并且当多于一个荧光团被成像时对于Ca 2+检测是最佳的。比率指示剂可以非常精确地校准，并且它们最大限度地减少了与化学Ca 2+指示剂相关的最常见问题，包括不均匀的染料负载、泄漏、光漂白和细胞体积的变化。最近的技术进步，允许在体内钙离子测量也将进行讨论。
Our understanding of the underlying mechanisms of Ca2+ signaling as well as our appreciation for its ubiquitous role in cellular processes and has been rapidly advanced, in large part, due to the development of fluorescent Ca2+ indicators. In this chapter, we discuss some of the most common chemical Ca2+ indicators that are widely used for the investigation of intracellular Ca2+ signaling. Advantages, limitations and relevant procedures will be presented for each dye including their spectral qualities, dissociation constants, chemical forms, loading methods and equipment for optimal imaging. Chemical indicators that are now available allow for intracellular Ca2+ detection over a very large range (<50 nM to >50 μM). Higher affinity indicators can be used to quantify Ca2+ levels in the cytosol while lower affinity indicators can be optimized for measuring Ca2+ in subcellular compartments with higher concentrations. Indicators can be classified into either single wavelength or ratiometric dyes. Both classes require specific lasers, filters, and/or detection methods that are dependent upon their spectral properties and both classes have advantages and limitations. Single wavelength indicators are generally very bright and optimal for Ca2+ detection when more than one fluorophore is being imaging. Ratiometric indicators can be calibrated very precisely and they minimize the most common problems associated with chemical Ca2+ indicators including uneven dye loading, leakage, photobleaching and changes in cell volume. Recent technical advances that permit in vivo Ca2+ measurements will also be discussed.
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