Ji Xiaoting, Wang Wei, Li Xiaoqian, Chen Yaoyao, Ding Caifeng
Key Laboratory of Sensor Analysis of Tumor Marker, Ministry of Education, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, PR China.
Key Laboratory of Sensor Analysis of Tumor Marker, Ministry of Education, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, PR China.
Talanta. 2016 Apr 1;150:666-70. doi: 10.1016/j.talanta.2016.01.004. Epub 2016 Jan 4.
In this paper, a new chemiluminescence (CL) strategy was constructed for the determination of physiological thiols by using an isoluminol labeled nano-probe. The amino group on the surface of the magnetic beads (MBs) were converted into pyridyl disulfide groups by treatment with N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP), in the meantime, isoluminol and thiolated signal DNA were labeled on the surface of the polystyrene microspheres (PSMs). By treating the SPDP activated MBs with the modified PSMs, isoluminol molecules on the surface of the PSMs, along with the thiolated signal DNA, were attached to the surface of the MBs through disulfide bonds to form a CL probe. In the presence of glutathione (GSH), the disulfide bonds could be cleaved readily. The isoluminol molecules modified on the surface of the PSMs released from the CL probes were detached by magnetic separation and transferred to the dark closet for CL detection of isoluminol-H2O2-HRP system. Using GSH detection as a model, we prove a linear dose response in the range from 5 × 10(-10) to 8 × 10(-8)M. The detection limit of this trial for GSH determined is 5 × 10(-10)M. The proposed design was successfully applied to the extracts of K562 cell for intracellular thiols detection, the average amount of thiols was about 4.114 × 10(-13)M per K562 cell.
本文构建了一种新的化学发光（CL）策略，用于使用异鲁米诺标记的纳米探针测定生理硫醇。通过用N-琥珀酰亚胺基-3-（2-吡啶二硫代）丙酸酯（SPDP）处理，将磁珠（MBs）表面的氨基转化为吡啶二硫基团，同时，将异鲁米诺和硫醇化信号DNA标记在聚苯乙烯微球（PSMs）表面。通过用修饰的PSMs处理SPDP活化的MBs，PSMs表面的异鲁米诺分子与硫醇化信号DNA一起通过二硫键连接到MBs表面，形成CL探针。在谷胱甘肽（GSH）存在下，二硫键可容易地裂解。从CL探针释放的PSMs表面修饰的异鲁米诺分子通过磁分离分离，并转移到暗室中用于异鲁米诺-H2O2-HRP系统的CL检测。以GSH检测为模型，我们证明在5×10（-10）至8×10（-8）M范围内呈线性剂量响应。该试验测定GSH的检测限为5×10（-10）M。所提出的设计成功应用于K562细胞提取物的细胞内硫醇检测，每个K562细胞中硫醇的平均量约为4.114×10（-13）M。
