Laserscanning Mikroscope

对于分子结构的空间分离和多种荧光染色信号的快速检测，需要一个共聚焦高分辨率显微镜系统。按照计划，该显微镜系统将由基尔大学生化研究所、药学研究所、生理学研究所、解剖学研究所以及合作研究中心877(SFB877)的成员使用。在CRC877的框架下，该系统将成为中央显微镜服务平台的一部分。对于活细胞成像，需要一个带有小室的倒置显微镜系统，允许温度调节和持续的气体供应。显微镜系统计划用于固定细胞、组织样本和活细胞。为了检测细胞通信的动态过程，需要以足够的记录速度同时检测不同的荧光信号。一个非常高的分辨率和快速的检测模式是必要的，允许由于低激光强度造成的最小的样品损失。为此，必须应用高像素驻留时间来成像活细胞模式中具有高灵敏度和低信噪比的即使是非常弱的信号。显微镜应允许常规成像以精确的空间和非重叠模式检测多色荧光信号。该系统的光学分辨率应该能够处理和成像非常微小的细胞内和细胞外结构、微环境的组成、共定位和荧光最近蛋白(FRET)的相互作用。这需要一种灵敏的检测方法，结合数学应用来分析信号和图像(例如去卷积)，以分别优化信号/噪声比和信号质量。为了对多个数据点、多个细胞和组织区域进行足够详细的成像，并在较长时间内以可重现的方式进行成像，需要一个机动扫描台。用于检测发射的波长应单独确定，并在任何时候再次调用。对于多标记信号的区分，发射需要详细的光谱分析。此外，不同的用户还需要用于相互作用研究、FRAP、光激活和使用灵敏荧光生物传感器进行生理学研究的标记定位的模块。为了监测活细胞模式中的动态过程，需要一种快速扫描系统来并行检测不同的标记。

A confocal high resolution microscope system is required for spatial separation of molecular structures and for the rapid detection of multiple fluorescence staining signals. It is planned that this microscope system will be used by members of the Biochemical Institute, the Institute of Pharmacy, Institute of Physiology, Institute of Anatomy and members of the collaborative research center 877 (SFB877) of the University Kiel. In the framework of the CRC877 the system will be part of the central microscopy service platform. For live cell imaging an inverse microscope system with a chamber is needed allowing temperature regulation and continous gas supply. The microscope system is planned for fixed cells, tissue samples and living cells. For the detection of dymanic processes of cell communication it is required to detect different fluorescence signals simultanously and with a sufficient speed of recording. A very high resolution and a rapid detection mode is necessary allowing a minumum of sample loss due to low laser intensities. For this purpose a high pixel dwell time to image even very weak signals with high sensitivity and low signal/noice ratio in the live cell modus has to be applied. The microscope should allow convocal imaging to detect multicolor fluorescence signals in a precise spatial and non overlapping mode. The optical resolution of the system should be able to process and image very tiny intracellular and extracellular structures, the composition of microenviroments, colocalisation and interaction of fluorecent proteins (FRET). This requires a sensitive detection methodology combined with mathematical applications to analyse signals and images (e.g. deconvolution) to optimize the signal/noice ratio and signal quality, respectively. To image multiple data points, multiple cells and tissue areas with sufficient detail and in a reproducible way over longer periods a motorized scanning table is needed. The wavelengths for the detection of the emission should be individually determined and called up again at any time. For the differentiation of signals of multilabelling the emission requires detailed spectral analysis. In addition moduls for interaction studies, FRAP, photo activation and the localisation of markers for physiological studies using sensitive fluorescent biosensors are required by different users. To monitor dynamic processes in a live cell modus a fast scanning system to detect in parallel different markers is required.