Mukherjee Kamalika, Chio Tak Ian, Gu Han, Banerjee Abhijit, Sorrentino Anthony M, Sackett Dan L, Bane Susan L
Department of Chemistry, Binghamton University, State University of New York , Binghamton, New York 13902, United States.
Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health , Bethesda, Maryland 20892, United States.
ACS Sens. 2017 Jan 27;2(1):128-134. doi: 10.1021/acssensors.6b00616. Epub 2017 Jan 18.
Detection and quantification of biomolecule carbonylation, a critical manifestation of oxidative stress, allows better understanding of associated disease states. Existing approaches for such analyses require further processing of cells and tissues, which leads to loss of both spatial and temporal information about carbonylated biomolecules in cells. Live cell detection of these species requires sensors that are nontoxic, sufficiently reactive with the biocarbonyl in the intracellular milieu, and detectable with commonly available instrumentation. Presented here is a new fluorescent sensor for biomolecule carbonyl detection: a hydrazine derivative of a benzocoumarin, 7-hydrazinyl-4-methyl-2H-benzo[h]chromen-2-one (BzCH), which meets these requirements. This probe is especially well suited for live cell studies. It can be excited by a laser line common to many fluorescence microscopes. The emission maximum of BzCH undergoes a substantial red shift upon hydrazone formation (from ∼430 to ∼550 nm), which is the result of fluorophore disaggregation. Additionally, the hydrazone exhibits an exceptionally large Stokes shift (∼195 nm). The latter properties eliminate self-quenching of the probe and the need to remove unreacted fluorophore for reliable carbonyl detection. Thus, biomolecule carbonylation can be detected and quantified in cells and in cell extracts in a one-step procedure using this probe.
生物分子羰基化是氧化应激的一种关键表现形式，对其进行检测和定量有助于更好地理解相关疾病状态。现有的此类分析方法需要对细胞和组织进行进一步处理，这会导致细胞中羰基化生物分子的空间和时间信息丢失。对这些物质进行活细胞检测需要无毒、能与细胞内环境中的生物羰基充分反应且能用常用仪器检测的传感器。本文介绍了一种用于生物分子羰基检测的新型荧光传感器：苯并香豆素的肼衍生物，7-肼基-4-甲基-2H-苯并[h]色烯-2-酮（BzCH），它满足这些要求。该探针特别适合活细胞研究。它可以被许多荧光显微镜常用的激光线激发。BzCH形成腙后发射最大值发生显著红移（从约430纳米到约550纳米），这是荧光团解聚的结果。此外，腙表现出异常大的斯托克斯位移（约195纳米）。后一种特性消除了探针的自猝灭现象，也无需去除未反应的荧光团即可进行可靠的羰基检测。因此，使用该探针可以在一步操作中对细胞和细胞提取物中的生物分子羰基化进行检测和定量。
