Chandran Neeli, Janardhanan Prajit, Bayal Manikanta, Unniyampurath Unnikrishnan, Pilankatta Rajendra, Nair Swapna S
Department of Physics, Central University of Kerala, Periye, Kasaragod, Kerala, India 671320.
Department of Biochemistry and Molecular Biology, Central University of Kerala, Periye, Kasaragod, Kerala, India 671320.
ACS Appl Bio Mater. 2020 Feb 17;3(2):1245-1257. doi: 10.1021/acsabm.9b01036. Epub 2020 Jan 30.
Metal nanoparticles-based sensors invoked much research attention in the biomedical field, especially in applications involving live cell imaging and monitoring. Here, a simple cost-effective method is adopted to synthesize glutathione coated copper nanoclusters (Cu-GSH NCs) with strong bright red fluorescence (625 nm). The clusters were found to be containing five Cu(0) atoms complexed with one molecule of glutathione (GSH) as evidenced by MALDI-TOF MS analysis. The synthesized Cu-GSH NCs system responds linearly to the pH in the acidic and alkaline ranges with a high degree of in vitro pH reversibility, projecting its potential as a real time pH sensor. Higher intensity emission observed in acidic conditions can be exploited for its employability as cellular organelle markers. The imaging and sensing potential of Cu-GSH NCs in the live human adenocarcinoma cell line, the HeLa cells, was tested. The treatment of HeLa cells for 48 h imparted deep red fluorescence, owing to the lower level of intracellular pH in cancer cells. In contrast, the imaging using normal cell lines (L-132, lung epithelial cell line) showed significantly lower fluorescence intensity as compared to that of HeLa cells. The subcellular pH-dependent fluorescence emission of Cu-GSH NCs was further assessed by treating HeLa cells with proton pump (V-ATPase) inhibitor Bafilomycin A1, which increases the vesicular pH. Interestingly, the fluorescent intensity of HeLa cells decreases with increasing concentration of Bafilomycin A1 in the presence of Cu-GSH NCs, as evidenced by the fluorescence microscopic images and quantitative fluorescent output. Accordingly, the developed Cu-GSH NCs system can be employed as an efficient pH-based bioimaging probe for the detection of cancer cells with an implied potential for the label free subcellular organelle tracking and marking. Importantly, the Cu-GSH NCs can be used for live cell pH imaging owing to their high degree of reversibility in sensing of pH variation.
基于金属纳米颗粒的传感器在生物医学领域引起了广泛的研究关注，尤其是在涉及活细胞成像和监测的应用中。在此，采用一种简单且经济高效的方法合成了具有强亮红色荧光（625 nm）的谷胱甘肽包覆铜纳米簇（Cu-GSH NCs）。基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）分析表明，这些纳米簇含有五个与一分子谷胱甘肽（GSH）络合的Cu(0)原子。合成的Cu-GSH NCs系统在酸性和碱性范围内对pH呈线性响应，具有高度的体外pH可逆性，显示出其作为实时pH传感器的潜力。在酸性条件下观察到的较高强度发射可用于其作为细胞器标记物的应用。测试了Cu-GSH NCs在人源腺癌活细胞系HeLa细胞中的成像和传感潜力。由于癌细胞内较低的pH水平，用Cu-GSH NCs处理HeLa细胞48小时后发出深红色荧光。相比之下，使用正常细胞系（L-132，肺上皮细胞系）进行的成像显示，与HeLa细胞相比，荧光强度明显较低。通过用质子泵（V-ATPase）抑制剂巴弗洛霉素A1处理HeLa细胞来进一步评估Cu-GSH NCs的亚细胞pH依赖性荧光发射，该抑制剂会增加囊泡pH。有趣的是，荧光显微镜图像和定量荧光输出表明，在存在Cu-GSH NCs的情况下，HeLa细胞的荧光强度随巴弗洛霉素A1浓度的增加而降低。因此，所开发的Cu-GSH NCs系统可作为一种基于pH的高效生物成像探针，用于检测癌细胞，具有无标记亚细胞细胞器追踪和标记的潜在应用。重要的是，由于Cu-GSH NCs在pH变化传感方面具有高度可逆性，可用于活细胞pH成像。
