3D-STED Microscope

小的亚细胞室的调查已显著改善了定位显微镜方法的使用。这些方法可以观察到分子信号复合物的结构和动态，是当今细胞生物学中的标准显微镜工具。定位精度比共聚焦显微镜提高了10倍左右，达到了电子显微镜的分辨率。主要的优点是探针的制备要简单得多，并且可以在活细胞中使用定位显微镜。随机方法，如光激活定位显微镜（PALM）或随机光学重建显微镜（STORM），在获得单个荧光团信息方面最明显，但依赖于许多图像的获取和精确的重建。这种被称为受激发射损耗显微镜（STED）的确定性方法使用一种确定的光模式来扫描样品，并允许更快地将分子定位相互关联。检测器和照明算法的优化提高了STED显微镜在活细胞中的可用性。自适应光学或光学元件校正的实现也允许在所有三个空间维度上具有几乎相似的定位精度。有了这些改进，STED显微镜现在适用于广泛的样品。随着STED显微镜的获得，我们希望为美因茨约翰内斯古腾堡大学生物系越来越多的分子/细胞生物学工作组提供一个有效的工具来进行他们的研究。在STED显微镜内使用脉冲激光还提供了使用荧光团的荧光活时间的可能性，并研究它们在相关维度上的相互作用。

Investigations of small subcellular compartments has been significantly improved by the use of localisation microscopy methods. These methods allow to look into the structure and dynamic of molecular signalling complexes and are today standard microscopy tools in cell biology. The gain of localisation precision is about tenfold in comparison to confocal microscopy and reach the resolution of electron microscopy. The main advantage is the much simpler preparation of the probes and the possibility to use localisation microscopy also in living cells. Stochastic approach as Photoactivation localisation microscopy (PALM) or Stochastic Optical Reconstruction Microscopy (STORM) is most obvious in the gain of single fluorophore information, but relay on the acquisition of many pictures and precise reconstructions. The deterministic approach, called Stimulated Emission Depletion microscopy (STED) use a defined light pattern to scan the sample and allow to correlate much faster the localisation of molecules to each other. The optimization of detectors and illumination algorithms has improved the usability of STED microscopy in living cells. Implementation of adaptive optics or corrections in the optical components allow also a nearly similar localisation precision in all three spatial dimensions. With these improvements the STED microscopy is now relevant for a broad spectrum of samples. With the acquisition of a STED microscope we like to provide a valid tool for the growing number of molecular/cell biological working groups in the biology department of the Johannes Gutenberg University Mainz to conduct their research. The use of pulsed lasers within the STED microscope offers in addition the possibility to use the fluorescent live time of the fluorophores and investigate their interactions in relevant dimensions to their local arrangement.