Fluorescence-intensity distribution analysis and its application in biomolecular detection technology
Fluorescence-intensity distribution analysis and its application in biomolecular detection technology
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DOI：
10.1073/pnas.96.24.13756
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发表时间：
1999-11-23
影响因子：
11.1
通讯作者：
Gall, K
中科院分区：
文献类型：
--
作者：
Kask, P;Palo, K;Gall, K
一种方法，荧光强度分布分析，已开发用于共聚焦显微镜研究，其中监测的荧光强度的样品与异质性的亮度分布。一个可调的公式，建模的空间亮度分布，和技术的生成函数的理论光子计数分布的计算作为两个基石的方法。该方法允许同时测定溶液中的单个荧光物质的浓度和特定亮度值。因此，我们提出了一个非常敏感的工具来监测荧光标记的分子或其他微粒与其各自的生物对应物的相互作用，应该在生命科学，医学和药物发现中找到广泛的应用。通过研究5 ′-（6-羧基四甲基罗丹明）-标记和未标记的互补寡核苷酸的杂交以及随后通过限制性内切酶对DNA杂交体的切割，证明了其潜力。
A methodology, fluorescence-intensity distribution analysis, has been developed for confocal microscopy studies in which the fluorescence intensity of a sample with a heterogeneous brightness profile is monitored. An adjustable formula, modeling the spatial brightness distribution, and the technique of generating functions for calculation of theoretical photon count number distributions serve as the two cornerstones of the methodology. The method permits the simultaneous determination of concentrations and specific brightness values of a number of individual fluorescent species in solution. Accordingly, we present an extremely sensitive tool to monitor the interaction of fluorescently labeled molecules or other microparticles with their respective biological counterparts that should find a wide application in life sciences, medicine, and drug discovery. Its potential is demonstrated by studying the hybridization of 5'-(6-carboxytetramethylrhodamine)-labeled and nonlabeled complementary oligonucleotides and the subsequent cleavage of the DNA hybrids by restriction enzymes.