Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province and ‡Experimental Chemistry Teaching Center, School of Chemistry and Chemical Engineering, Shaanxi Normal University , 620 Xi Chang'an Street, Xi'an, Shaanxi 710119, People's Republic of China.
Anal Chem. 2015 Nov 17;87(22):11332-6. doi: 10.1021/acs.analchem.5b02612. Epub 2015 Nov 5.
Click chemistry with metabolic labeling has been widely used for selectively imaging biomacromolecules in cells. The first example of azide-alkyne cycloaddition for ratiometric fluorescent imaging of live cells is reported. The precursor of the azido fluorophore (cresyl violet) has a fluorescence emission peak at 620 nm. The electron-rich nitrogen of the azido group blue-shifts the emission peak to 566 nm. When the click reaction occurs, an emission peak appears at 620 nm due to the lower electronic density of the newly formed triazole ring, which allows us to ratiometrically record fluorescence signals. This emission shift was applied to ratiometric imaging of propargylcholine- and dibenzocyclooctyne-labeled human breast cancer cells MCF-7 under laser confocal microscopy. Two typical triazole compounds were isolated for photophysical parameter measurements. The emission spectra presented a fluorescence emission peak around 620 nm for both click products. The results further confirmed the emission wavelength change was the result of azide-alkyne cycloaddition reaction. Since nearly all biomolecules can be metabolically labeled by reported alkyne-functionalized derivatives of native metabolites, our method can be readily applied to image these biomacromolecules.
点击化学与代谢标记已被广泛用于选择性地对细胞内的生物大分子进行成像。本文报道了首例用于活细胞比率荧光成像的叠氮-炔基环加成反应的实例。叠氮荧光团的前体（甲酚紫）的荧光发射峰在 620nm 处。叠氮基团富电子的氮原子使发射峰蓝移至 566nm。当点击反应发生时，由于新形成的三唑环的电子密度降低，会出现一个在 620nm 处的发射峰，这使我们能够进行比率记录荧光信号。这种发射位移被应用于激光共聚焦显微镜下对丙炔胆碱和二苯并环辛炔标记的人乳腺癌细胞 MCF-7 的比率成像。两种典型的三唑化合物被分离出来进行光物理参数测量。对于点击产物，其发射光谱均在 620nm 左右呈现出荧光发射峰。该结果进一步证实了发射波长的变化是叠氮-炔基环加成反应的结果。由于几乎所有的生物分子都可以被报道的天然代谢物的炔基功能化衍生物进行代谢标记，因此我们的方法可以很容易地应用于这些生物大分子的成像。
