Uptake, distribution and diffusivity of reactive fluorophores in cells: implications toward target identification.
Uptake, distribution and diffusivity of reactive fluorophores in cells: implications toward target identification.
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DOI：
10.1021/mp100089k
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发表时间：
2010-08-02
影响因子：
4.9
通讯作者：
Krise JP
中科院分区：
文献类型：
--
作者：
Cunningham CW;Mukhopadhyay A;Lushington GH;Blagg BS;Prisinzano TE;Krise JP
近年来，人们对无铜点击化学在体内和体外研究广泛的生物学事件的应用产生了很大兴趣。具体地，叠氮化物缀合的荧光探针可用于鉴定已经用生物正交反应基团修饰的靶标。对于这种化学的细胞内应用，荧光叠氮化物的结构和物理化学性质变得越来越重要。理想的荧光团应该在细胞内广泛积累，具有均匀的细胞内分布，并且是自由的（未结合的），使它们能够有效地参与双分子反应。我们在这里报告的合成和评估一组结构不同的荧光探针，以检查其潜在的有用性在细胞内点击反应。使用定量和定性方法比较评估总细胞摄取和细胞内分布概况。使用基于光漂白后荧光恢复（FRAP）的方法测量细胞内扩散系数。许多反应性荧光团表现出次优性能的细胞内反应。基于BODIPY和TAMRA的叠氮化物具有上级的细胞积累，而基于TAMRA的探针具有最均匀的细胞内分布和最佳的细胞溶质扩散性。总的来说，这些结果提供了一个公正的比较评价叠氮化物连接的荧光团细胞内点击反应的适用性。
There is much recent interest in the application of copper-free click chemistry to study a wide range of biological events in vivo and in vitro. Specifically, azide-conjugated fluorescent probes can be used to identify targets which have been modified with bioorthogonal reactive groups. For intracellular applications of this chemistry, the structural and physicochemical properties of the fluorescent azide become increasingly important. Ideal fluorophores should extensively accumulate within cells, have even intracellular distribution, and be free (unbound), allowing them to efficiently participate in bimolecular reactions. We report here on the synthesis and evaluation a set of structurally diverse fluorescent probes to examine their potential usefulness in intracellular click reactions. Total cellular uptake and intracellular distribution profiles were comparatively assessed using both quantitative and qualitative approaches. The intracellular diffusion coefficients were measured using a fluorescence recovery after photobleaching (FRAP)-based method. Many reactive fluorophores exhibited suboptimal properties for intracellular reactions. BODIPY- and TAMRA-based azides had superior cellular accumulation, whereas TAMRA-based probes had the most uniform intracellular distribution and best cytosolic diffusivity. Collectively, these results provide an unbiased comparative evaluation regarding the suitability of azide-linked fluorophores for intracellular click reactions.
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DOI：
10.1073/pnas.0707090104
发表时间：
2007-10-23
影响因子：
11.1
作者：
Baskin, Jeremy M.;Prescher, Jennifer A.;Bertozzi, Carolyn R.
通讯作者：
Bertozzi, Carolyn R.
影响因子：
4.8
作者：
Duvvuri, M;Gong, YP;Krise, JP
通讯作者：
Krise, JP
影响因子：
4
作者：
Duvvuri, Muralikrishna;Konkar, Saimidha;Krise, Jeffrey P.
通讯作者：
Krise, Jeffrey P.
影响因子：
5.2
作者：
Mottram, Laurie F.;Maddox, Ewa;Peterson, Blake R.
通讯作者：
Peterson, Blake R.
影响因子：
4
作者：
Laughlin, Scott T.;Bertozzi, Carolyn R.
通讯作者：
Bertozzi, Carolyn R.