EuroTracker Dyes: Synthesis and Application in Functional Cell Imaging

通过光学显微镜观察某些活细胞或微生物需要引入明亮的发光染料或染色剂，这些染料或染色剂可以有效地照亮细胞的选定部分。我们将开发稀土元素铕（EuroTracker染料）的明亮配合物，作为光学探针，有效地发光，并向观察者报告有关其化学或生物环境的信息。这些探针被设计成允许它们被物理科学家、生物化学家和细胞生物学家使用的显微镜中可用的光源激发，以研究细胞或相关小隔间的内部组成。通过检查一系列密切相关的探针，我们将定义定位于细胞特定隔室的系统，使其能够选择性地可视化。目前使用的光学探针要么基于有机化合物，要么基于大型人造蛋白质，要么基于所谓的量子点；每一种探针都存在化学稳定性低、容易褪色的问题，在某些情况下，它们对活细胞有相当大的毒性。因此，我们的目标是改变显微镜界的想法，并为这种发射金属配合物在生物学中用作染色剂和探针铺平道路。与传统探针相比，这些金属复合探针在更长的时间尺度上发光，因此在脉冲激发后，它们有更多的时间来收集发射的光，而用于激发它们的光的曝光时间很短。较长的发射寿命避免了与光散射和背景自身荧光相关的数据采集问题。同时，我们将开发仪器方法，使这些探测器的光谱特征能够使用快速成像方法快速记录。为此目的，我们将与显微镜制造商和光学专家合作。这将允许在10秒的时间尺度上跟踪细胞内化学成分的变化。作为第一步，我们将开发出能够报告pH值变化以及特定细胞区室内钠、镁和锌含量的探针，从而实时跟踪活细胞中这些重要金属离子含量的变化。通过采用模块化方法来设计探针，随后可以通过在探针结构中引入适当的金属结合部分来检测不同的金属离子。

The visualisation of certain living cells or microbes by optical microscopy requires the introduction of bright, luminescent dyes or stains, that can light up selected parts of the cell effectively.We will develop bright complexes of the rare earth element, europium, (EuroTracker dyes) as optical probes that emit light efficiently and report information back to the observer about their chemical or biological environment. These probes have been designed to allow them to be excited by the light sources that are available in microscopes used by physical scientists, biochemists and cell biologists, to study the interior composition of cells or related small compartments. By examining series of closely related probes, we shall define systems that locate to a particular compartment of a cell, allowing it to be visualised selectively. The optical probes that are currently used are based either on organic compounds, on large man-made proteins or on so-called quantum dots; each of these probes suffer from problems of low chemical stability, a tendency to fade and, in certain cases, they are rather toxic to the living cell. Thus we aim to change the thinking of the microscopy community and to pave the way for such emissive metal complexes to be used as stains and probes in biology. These metal complex probes emit light over longer timescales than conventional probes, and so their is more time to collect the emitted light after pulsed excitation and only short exposure times to the light that is used to excite them. The long emission lifetime avoids problems in data acquisition that are associated with light scattering and background autofluorescence. In parallel, we shall develop instrumentation methods that allow the spectral signature of these probes to be recorded quickly using fast imaging methods. We shall work with microscope manufacturers and with optics specialists to this end. This will allow changes in chemical composition within a cell to be tracked on a 10 second timescale. As a first step, we shall develop probes that can report on pH change and on the amount of sodium, magnesium and zinc within a specific cell compartment, allowing changes in the amount of these important metal ions to be tracked in living cells in real time. By adopting a modular approach to probe design, different metal ions can be examined subsequently, by introducing the appropriate metal-binding moiety into the probe structure.