Bioorthogonal turn-on probes for imaging small molecules inside living cells.
Bioorthogonal turn-on probes for imaging small molecules inside living cells.
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DOI：
10.1002/anie.200906120
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发表时间：
2010-04-06
影响因子：
16.6
通讯作者：
Weissleder, Ralph
中科院分区：
文献类型：
--
作者：
Devaraj, Neal K.;Hilderbrand, Scott;Upadhyay, Rabi;Mazitschek, Ralph;Weissleder, Ralph
Bioorthogonal “click” chemistries are now widely used in chemical biology for a myriad of applications such as activity based protein profiling, monitoring cell proliferation, generation of novel enzyme inhibitors, monitoring the synthesis of newly formed proteins, protein target identification, and studying glycan processing.[1, 2] Arguably, the most fascinating applications involve using these bioorthogonal chemistries to assemble molecules in the presence of living systems such as live cells or even whole organisms.[3, 4] These latter applications require that the chemistry does not employ toxic metal catalysts and maintains kinetics that allow fast reaction to occur with micromolar concentrations of reagents in a time span of minutes to hours. To fulfill these criteria, various “copper-free” click chemistries have been reported, such as the strain-promoted azide-alkyne cycloaddition and the Staudinger ligation, to react with azides on the surface of live cells both in culture and in in vivo systems such as mice and zebrafish.[4] However, to date, the application of “click” chemistries in living systems, has been largely limited to extracellular targets.[5] The reasons for this are likely several. In addition to fulfilling the stability, toxicity, and chemoselectivity requirements of “click” chemistry, intracellular live cell labeling requires reagents that can pass easily through biological membranes and kinetics that enable rapid labeling even with the low concentrations of agent that make it across the cell membrane. Additionally, a practical intracellular bioorthogonal coupling scheme would need to incorporate a mechanism by which the fluorescent tag increases in fluorescence upon covalent reaction to avoid visualizing accumulated but unreacted imaging agent (ie" background"). Such activatable “turn-on” probes would significantly increase the signal-tobackground ratio, which is particularly relevant to imaging targets inside living cells since a stringent washout of unreacted probe is not possible.In previous years a number of elegant probes have been introduced whose fluorescence increases after azide-alkyne cycloaddition or staudinger ligation coupling reactions.[6] Most of these strategies utilize a reactive group intimately attached to the fluorophore thus necessitating synthesis of new fluorophore scaffolds or take advantage of a FRET based activation requiring appending of an additional molecule that can act as an energy transfer agent. Furthermore, most probes employing these popular coupling schemes have not been used to label intracellular targets in live cells. Here we report a series of activatable “turnon” tetrazine-linked fluorescent probes, which react rapidly via an inverse electron demand cycloaddition with strained dienophiles such as trans-cyclooctene. Upon cycloaddition, the fluorescence intensity increases dramatically, in some cases by~ 20 fold. This fluorescence
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影响因子：
15
作者：
Blackman, Melissa L.;Royzen, Maksim;Fox, Joseph M.
通讯作者：
Fox, Joseph M.
影响因子：
16.6
作者：
Devaraj, Neal K.;Upadhyay, Rabi;Hatin, Jered B.;Hilderbrand, Scott A.;Weissleder, Ralph
通讯作者：
Weissleder, Ralph
影响因子：
16.6
作者：
Ning, Xinghai;Guo, Jun;Boons, Geert-Jan
通讯作者：
Boons, Geert-Jan
影响因子：
4.8
作者：
Farinas, J;Verkman, AS
通讯作者：
Verkman, AS
DOI：
10.1083/jcb.94.3.688
发表时间：
1982-09
期刊：
The Journal of cell biology
影响因子：
--
作者：
Manfredi JJ;Parness J;Horwitz SB
通讯作者：
Horwitz SB