一种含聚乙二醇非离子亲水基团的新型水溶性荧光染料的合成及细胞成像

通过向吲哚环“N”位上引入一个含聚乙二醇(PEG)醚链的非离子亲水基团,合成了一种新型水溶性不对称五甲川吲哚菁染料。 用核磁(¹H NMR)和高分辨质谱(HRMS)表征染料的结构,测试了染料的光谱性能,标记牛血清白蛋白(BSA),并对固定细胞和活细胞分别染色。 结果表明,该染料在水中的最大紫外吸收波长和荧光发射波长分别为648和668 nm,斯托克斯(stokes)位移为20 nm,荧光量子产率(Ф)为0.13,以碘钨灯为光源光照8 h后,染料光降解率为5.8%。 用染料的NHS活性酯标记牛血清白蛋白,标示率(D/P)为1.16。 对固定细胞染色发现,染料可对细胞整体着色,对细胞核染色最明显,能清晰看到核仁。 对活细胞染色发现,有少量染料跨膜进入细胞内部,对细胞质和细胞核有微弱染色,但在活细胞膜上聚集明显。

Synthesis and Cell markers of Novel Water-soluble Asymmetric Fluorescent Dyes with Nonionic Hydrophilic Group

In this work, a novel asymmetric pentamethine cyanine(Cy5) dye containing one nonionic hydrophilic group with two polyethylene glycol(PEG) ether chains on the indoline “N” was synthesized and characterized by ¹H nuclear magnetic resonance(¹H NMR) and high resolution mass spectrometer(HRMS). The spectra character and light stability of this Cy5 were investigated. The results show that the λ_ab and λ_em of the dye in water are 648 nm and 668 nm, respectively, and Stokes shifts reach 20 nm. The fluorescence quantum yield(Ф) in water is 0.13. After irradiated for 8 h, the photo-degradation of the aqueous soluble dyestuff of 5.8% happens. Albumin Bovine Serum(BSA) is labeled with this Cy5, and the dye/protein molar ratio(D/P) of fluorescent labeling reachs 1.16. By comparing fixed cells staining and viable cells staining, it is found that fixed cells can be overall stained by the new Cy5, the nucleoli of which are the most obvious and can be seen clearly, and that a small amount of the new Cy5 across the cell membrane of viable cells into the cell can stain cytoplasm and cell nucleus weakly, but a lot of the dye clumps together in the viable cell membrane.