Photoactivatable Fluorophores for Super-Resolution Imaging Based on Oxazine Auxochromes
Photoactivatable Fluorophores for Super-Resolution Imaging Based on Oxazine Auxochromes
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DOI：
10.1021/jp211796p
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发表时间：
2012-03-15
影响因子：
3.7
通讯作者：
Raymo, Francisco M.
中科院分区：
文献类型：
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作者：
Deniz, Erhan;Tomasulo, Massimiliano;Raymo, Francisco M.
我们合成了五个荧光团-光致变色二联体，其设计在光学控制下在非荧光和荧光异构体之间可逆地切换。这些化合物在其分子骨架中将恶嗪光致变色剂与联苯、芴、芘、香豆素或花青荧光团配对，并且可以从已知的前体以30至63%的产率在单个步骤中制备。核磁共振光谱表明，这些化合物的恶嗪环打开和关闭在毫秒级的时间尺度在乙腈中在环境温度下自发。在这些条件下，平衡时开环异构体的分数在所有情况下都可以忽略不计，除了花青衍生物，它几乎完全是这种形式。然而，吸收和发射光谱表明，开环异构体的分数是敏感的溶剂极性和增加的过渡从乙腈到甲醇。或者，开环异构体可以光化学填充或通过添加酸捕获。在这两种情况下，特征吸收和发射带的3 H-吲哚生色团，嵌入在开环物种，可以清楚地观察到在可见光区。在香豆素衍生物的情况下，该发色团片段的亮度足够高以允许以优异的信噪比对单个分子成像。事实上，单荧光团-光致变色二联体的荧光可以在紫外输入的影响下被激活，并且所产生的物种可以在可见光照射下以纳米级精度定位。事实上，亚衍射图像的聚合物纳米粒子，掺杂这种特殊的二分体，可以重建与纳米级的分辨率。因此，我们在单分子水平上进行荧光切换的操作原理可以提供克服衍射的机会，并最终导致开发用于超分辨率荧光成像的整个系列探针。
We synthesized five fluorophore-photochrome dyads designed to switch reversibly between nonfluorescent and fluorescent isomers under optical control. These compounds pair an oxazine photochrome to a biphenyl, fluorene, pyrene, coumarin, or cyanine fluorophore in their molecular skeleton and can be prepared in a single step from known precursors in yields ranging from 30 to 63%. Nuclear magnetic resonance spectroscopy indicates that the oxazine ring of these compounds opens and closes spontaneously on a millisecond time scale in acetonitrile at ambient temperature. Under these conditions, the fraction of ring-open isomer at equilibrium is negligible in all instances with the exception of the cyanine derivative, which instead is almost exclusively in this form. Absorption and emission spectroscopies demonstrate, however, that the fraction of ring-open isomer is sensitive to solvent polarity and increases with a transition from acetonitrile to methanol. Alternatively, the ring-open isomer can be populated photochemically or trapped with the addition of acid. In both instances, the characteristic absorption and emission bands of the 3H-indolium chromophores, embedded within the ring-open species, can clearly be observed in the visible region. In the case of the coumarin derivative, the brightness of this chromophoric fragment is sufficiently high to permit the imaging of individual molecules with excellent signal-to-noise ratios. In fact, the fluorescence of single fluorophore-photochrome dyads can be activated under the influence of ultraviolet inputs and the resulting species can be localized with nanoscale precision under visible illumination. Indeed, subdiffraction images of polymer nanoparticles, doped with this particular dyad, can be reconstructed with nanoscale resolution. Thus, our operating principles for fluorescence switching at the single-molecule level can offer the opportunity to overcome diffraction and, eventually, lead to the development of an entire family of probes for super-resolution fluorescence imaging.