Institute of Environmental Science, College of Chemistry and Chemical Engineering, Shanxi University, Taiyuan, 030006, P. R. China.
Translational medicine research center, Shanxi Medical University, Taiyuan, 030006, P. R. China.
Analyst. 2019 Jan 21;144(2):439-447. doi: 10.1039/c8an01908h. Epub 2018 Nov 13.
We report herein a two-photon ratiometric fluorescent probe (DNEPI) for mitochondrial cysteine (Cys) detection on the basis of a merocyanine (compound 1) as the two-photon fluorophore and a 2,4-dinitrobenzensulfonyl (DNBS) unit as the biothiol reaction site. Upon reaction with Cys in DMSO/PBS (1/1, v/v), DNEPI showed a distinct ratiometric fluorescence emission characteristic (F/F) linearly proportional to Cys concentrations over the range of 2-10 μM, which was attribute to the enhanced intramolecular charge transfer (ICT) effect by cleavage of the sulfonic acid ester bond of DNEPI to release compound 1. More importantly, the probe could detect Cys with a fast response time (within 2 min) and the detection limit was quantitatively calculated as 0.29 μM. Furthermore, DNEPI not only exhibited high selectivity toward Cys over other similar biothiols, including homocysteine (Hcy) and glutathione (GSH), but also displayed significant mitochondrial-targeting ability, which were favorable for mitochondrial Cys-selective imaging. Subsequently, application of DNEPI to Cys imaging in live cells was successfully achieved by two-photon fluorescence microscopy, suggesting that the probe proposed here could be used to monitor mitochondrial Cys concentration changes in live cells with negligible interference from other biological thiols.
我们基于甲川菁（化合物 1）作为双光子荧光团和 2,4-二硝基苯磺酰（DNBS）单元作为生物硫醇反应位点，报道了一种用于检测线粒体半胱氨酸（Cys）的双光子比率荧光探针（DNEPI）。在 DMSO/PBS（1/1，v/v）中与 Cys 反应时，DNEPI 表现出明显的双光子荧光发射特征（F/F），与 Cys 浓度呈线性比例，范围为 2-10 μM，这归因于 DNEPI 的磺酸酯键断裂，释放出化合物 1，从而增强了分子内电荷转移（ICT）效应。更重要的是，该探针能够快速响应（在 2 分钟内）检测 Cys，检测限定量计算为 0.29 μM。此外，DNEPI 不仅对 Cys 具有较高的选择性，对其他类似生物硫醇（包括同型半胱氨酸（Hcy）和谷胱甘肽（GSH））具有较高的选择性，而且表现出显著的线粒体靶向能力，这有利于线粒体 Cys 选择性成像。随后，通过双光子荧光显微镜成功地将 DNEPI 应用于活细胞中的 Cys 成像，表明这里提出的探针可以用于监测活细胞中线粒体 Cys 浓度的变化，几乎没有其他生物硫醇的干扰。
