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首页> 外文会议>Ultrasensitive and Single-Molecule Detection Technologies II; Progress in Biomedical Optics and Imaging; vol.8 no.21; Proceedings of SPIE-The International Society for Optical Engineering; vol.6444 >Single-molecule studies on individual metal complexes
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Single-molecule studies on individual metal complexes

机译：单个金属配合物的单分子研究

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Some fluorescent dyes, like Atto635 or Cy5, exhibit alterations in fluorescence emission in the presence of various metal ions and metal ion complexes. We used this effect to design dye-ligand conjugates that can be immobilized on glass surfaces and allow studying metal-ion binding using time-resolved single-molecule fluorescence spectroscopy (SMFS). Double-stranded DNA served as a rigid scaffold carrying 2,2'-bipyridene-4,4'-dicarboxylic acid as chelating ligand and a fluorescent dye as reporter, placed in close vicinity to the ligand. In the absence of metal ions, the probes showed high fluorescence quantum yield, whereas strong fluorescence quenching upon binding of Cu~(2+)-ions was observed. Time-resolved single-molecule measurements revealed stochastic switching between a highly fluorescent ("on") and a low fluorescent ("off") state. The coordination of the metal ion to the ligand is thus indicated by intramolecular fluorescence quenching of the dye. We screened various fluorescent dyes for their sensitivity to Cu~(2+)-coordination, and found that both Atto620 and MR121 are well-suited for this application. Ensemble studies of the fluorescence lifetimes of metal-sensors with Atto620 showed only small dependence on the metal-ion concentration, while single-molecule studies reported strong changes in the fluorescence lifetimes which were correlated with the observed on- and off-states. Our results further indicate that the fluorescence of Atto620 is not completely quenched upon association of the metal-ion complex; either because a less fluorescent complex is formed or because of intramolecular collisional quenching due to conformational changes of the C6-linker used for covalent coupling of the fluorescent dye.
机译：在存在各种金属离子和金属离子络合物的情况下，某些荧光染料（如Atto635或Cy5）在荧光发射方面表现出变化。我们利用这种效应设计了可以固定在玻璃表面上的染料-配体共轭物，并允许使用时间分辨的单分子荧光光谱法（SMFS）研究金属离子的结合。双链DNA充当刚性支架，携带2,2'-联吡啶-4,4'-二羧酸作为螯合配体，荧光染料作为报告分子，紧靠配体放置。在不存在金属离子的情况下，探针显示出高的荧光量子产率，而在结合Cu〜（2 +）-离子时观察到强荧光猝灭。时间分辨的单分子测量揭示了在高荧光（“开”）状态和低荧光（“关”）状态之间的随机切换。因此，通过染料的分子内荧光猝灭来指示金属离子与配体的配位。我们筛选了各种荧光染料对Cu〜（2 +）-配位的敏感性，发现Atto620和MR121都非常适合此应用。使用Atto620对金属传感器的荧光寿命进行的综合研究表明，对金属离子浓度的依赖性很小，而单分子研究则报告了荧光寿命的强变化，这与观察到的通态和关态有关。我们的结果进一步表明，Atto620的荧光在金属离子络合物缔合后并未完全被淬灭。要么是因为形成的荧光复合物较少，要么是由于用于荧光染料共价偶联的C6-接头构象变化导致分子内碰撞猝灭。

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