College of Chemistry and Chemical Engineering, Central South University, Changsha, Hunan 410083, PR China.
College of Chemistry and Chemical Engineering, Central South University, Changsha, Hunan 410083, PR China.
Talanta. 2017 Dec 1;175:470-476. doi: 10.1016/j.talanta.2017.07.071. Epub 2017 Jul 25.
Recently, membrane proteins have been considered as candidate cancer biomarkers and drug targets, due to their important roles in numerous physiological processes. Therefore, a facile, sensitive and quantitative detection of the membrane proteins is crucial for better understanding their roles in cancer cells and further validating their function in clinical research. We report a highly facile and sensitive detection method for membrane proteins on living cells in situ based on membrane protein-triggered release of cytosine (C)-rich single-stranded DNA (ssDNA) sequences, and the subsequent silver nanoclusters (AgNCs) transfer from polymer to C-rich ssDNA. The high-quantum yield and stable DNA-AgNCs allow the accurate detection of membrane proteins with facile operations and a common fluorescence spectrophotometer. The detection of protein tyrosine kinase-7 (PTK7), a membrane protein model, displays a response range from 30pM to 2nM with a detection limit of 12pM. The expression of PTK7 on single Hela cell and CCRF-CEM cell was calculated to be 7.5 × 10mol and 1.8 × 10mol, respectively. Given the simple and facile operation of this method, this detection platform can be applied as a universal strategy for ultrasensitive detection of membrane protein on cell in situ.
最近，由于膜蛋白在许多生理过程中起着重要作用，因此它们被认为是癌症生物标志物和药物靶点的候选物。因此，对膜蛋白进行简便、灵敏和定量的检测对于更好地了解它们在癌细胞中的作用以及进一步验证它们在临床研究中的功能至关重要。我们报道了一种基于膜蛋白触发释放富含胞嘧啶（C）的单链 DNA（ssDNA）序列的简便、灵敏的活细胞内原位膜蛋白检测方法，以及随后的聚合物到富含 C 的 ssDNA 的银纳米簇（AgNCs）转移。高量子产率和稳定的 DNA-AgNCs 允许使用简便的操作和普通荧光分光光度计准确检测膜蛋白。以膜蛋白模型蛋白酪氨酸激酶 7（PTK7）的检测为例，其响应范围从 30pM 到 2nM，检测限为 12pM。单个 Hela 细胞和 CCRF-CEM 细胞上 PTK7 的表达分别计算为 7.5×10-16mol 和 1.8×10-16mol。鉴于该方法操作简单，这种检测平台可作为一种通用策略，用于活细胞内原位膜蛋白的超灵敏检测。
