Generation of luminescent materials through living micoalgae

在这个后续的项目中，我们想进一步研究生物矿化过程中的海洋Emiliania huxleyi。我们希望揭示稀土铽从周围介质进入细胞内球石囊泡（cv）的途径，在那里产生球石。在该项目的第1部分中证明了Tb 3+掺入到球石中，这些结果表明，Tb 3+掺入到细胞内cv中生长的方解石球石中。研究Tb 3+通过细胞的途径的优点是当用光激发时其特异性荧光。为了研究Tb 3+在细胞中的转运和途径，我们想使用两种不同的方法：（A），我们想使用2D电泳鉴定参与Tb通过细胞转运的蛋白质。在这里，我们想比较钙化E。huxleyi菌株CCMP 1516与非钙化菌株CCMP 1516 NC以及培养基中不同量的Tb 3+和/或Ca 2+的比较。(B)我们希望通过共聚焦显微镜和电子显微镜技术揭示Tb从细胞膜到cv的过程中在细胞的哪些细胞器和/或隔室中积累。首先，我们将使用共聚焦显微镜观察在培养基中添加Tb 3+后的活细胞。我们希望使用滤波器跟踪Tb 3+在细胞中的分布，从而我们可以专门观察Tb 3+发射。由于细胞内球石的形成发生在1小时至2小时内，我们将评估在此期间细胞内Tb 3+的积累。当我们可以观察到其在细胞中特定部位的积累时，将对相应的细胞进行采样并在冷冻条件下通过TEM进行进一步处理，即首先用高压冷冻机（HPF）冷冻细胞，冷冻替代它们，并在切割后使用Cryo TEM/EDX和配备CL的TEM进行分析。

In this follow-up project we want to further investigate biomineralization processes in the marine alga Emiliania huxleyi. We want to reveal the pathway of the lanthanide terbium from the surrounding medium into the intracellular coccolith vesicle (cv), where the coccolith is produced. The incorporation of Tb3+ into the coccoliths was proven in part 1 of the project, these results showed, that Tb3+ is incorporated into the growing calcite coccolith in the cv inside the cells. The advantage of investigating the pathway of Tb3+ through the cells is its specific fluorescence when excited with light. For the investigation of the transport and pathway of Tb3+ in the cell, we want to use two different approaches: (A), we want to identify the proteins which are involved in the transportation of Tb through the cell using 2D electrophoresis. Here we want to compare the proteomes of the calcifying E. huxleyi strain CCMP1516 with the non-calcifying strain CCMP1516NC and at different amounts of Tb3+ and/or Ca2+ in the medium. (B), we want to reveal in which organelles and/or compartments of the cells the Tb is accumulated during its way from the cell membrane to the cv by Confocal microscopy and Electron microscopy techniques. First, we will observe the living cells after adding Tb3+ to the medium using a confocal microscope. We want to track the distribution of the Tb3+ in the cells using a filter, thereby we can visualize exclusively the Tb3+ emission. Since the formation of the intracellular coccoliths occurs within 1h to 2h, we will evaluate, where inside the cells the Tb3+ is accumulated during this time period. When we can observe its accumulation at a specific site in the cells, the corresponding cells will be sampled and further treated for the investigation by TEM under cryo-conditions, i.e. first freezing the cells with high pressure freezer (HPF), cryo-substitute them and after cutting analyze them using Cryo TEM/EDX and a TEM equipped with CL.