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Using diaminofluoresceins (DAFs) in nitric oxide research
Nitric Oxide ( IF 3.9 ) Pub Date : 2021-07-26 , DOI: 10.1016/j.niox.2021.07.002
Junjie Li 1 , Anthea LoBue 1 , Sophia K Heuser 1 , Francesca Leo 1 , Miriam M Cortese-Krott 1
Affiliation  

Diaminofluoresceins (DAFs) are fluorescent probes widely applied to measure nitric oxide (NO) formation in cells and tissues. The main advantages of these compounds are their availability and low cost, and the general availability of instruments able to detect green fluorescence in all laboratories; these include fluorimeters, flow cytometers, and fluorescent microscopes. What made these molecules particularly interesting for many scientists approaching the NO field is that they are apparently very easy to use, as compared with other techniques requiring specific instrumentation and knowledge like chemiluminescence and electron paramagnetic resonance. However, the reactivity and biological chemistry of these probes in the cellular environment is rather complex and still not fully understood. Moreover, secondary reactions with ascorbate, or interference with thiols occur in cells. Therefore, the use of DAFs requires specific experimental planning and a careful interpretation of the results obtained. In this methodological review, we described in detail what is known about the reactivity of DAFs, their application in biological assays, list some principles to help experimental planning, including the necessary controls, and list the caveats concerning result interpretation. These guiding principles will help to understand the “Method behind our DAF-madness”.



中文翻译:

在一氧化氮研究中使用二氨基荧光素 (DAF)

二氨基荧光素 (DAF) 是广泛应用于测量细胞和组织中一氧化氮 (NO) 形成的荧光探针。这些化合物的主要优点是它们的可用性和低成本,以及能够在所有实验室检测绿色荧光的仪器的普遍可用性;这些包括荧光计、流式细胞仪和荧光显微镜。使这些分子对许多接近 NO 领域的科学家特别感兴趣的是,与其他需要特定仪器和知识(如化学发光和电子顺磁共振)的技术相比,它们显然非常易于使用。然而,这些探针在细胞环境中的反应性和生物化学相当复杂,仍未完全了解。此外,与抗坏血酸的二次反应,或干扰硫醇发生在细胞中。因此,使用 DAF 需要具体的实验计划和对获得的结果的仔细解释。在本方法回顾中,我们详细描述了 DAF 的反应性及其在生物测定中的应用,列出了一些有助于实验计划的原则,包括必要的控制,并列出了有关结果解释的注意事项。这些指导原则将有助于理解“ 包括必要的控制,并列出有关结果解释的注意事项。这些指导原则将有助于理解“ 包括必要的控制,并列出有关结果解释的注意事项。这些指导原则将有助于理解“我们疯狂 DAF 背后的方法”。

更新日期:2021-08-05
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