Fluorescence/patch clamp setup

荧光/膜片钳设置

• 批准号: 438033605
• 金额: --
• 依托单位: Albert-Ludwigs-Universität Freiburg
• 依托单位国家: 德国
• 项目类别: Major Research Instrumentation
• 财政年份: 2020
• 资助国家: 德国
• 起止时间: 2019-12-31 至 无数据
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/438033605?language=en
• 关键词: Fluorescence; patch; clamp; setup

We apply for a fluorescence-patch clamp combination, which will be mainly used for infection work with human pathogens of risk group 2. This device ensemble that will be unique at the University of Freiburg is essential for the future research activities of the working group of Winfried Römer in order to be able to continue to conduct top research.The Römer lab explores the molecular mechanisms of the interactions of bacteria with various mammalian cells, e.g. lung, skin and immune cells. Here, the focus is on bacterial lectins, which recognize glycosylated receptors on the host cell plasma membrane and induce diverse signalling pathways that influence the physiology of the cell - in both a positive and a negative sense. The strength of our approach lies in the fact that, on the one hand, coming from complex cell biology, we can identify key molecules, and with this knowledge we can rebuild natural processes in simpler, synthetic systems and thus better understand them. Many important receptors, e.g. growth factor receptors, cell adhesion molecules, glycosphingolipids, and ion channels are targets of these bacterial lectins that massively affect the organization, dynamics, and stability of the plasma membrane.We had to realize that one technique alone, e.g. fluorescence microscopy, is by far not sufficient to adequately describe the complexity of cellular processes. With the help of the fluorescence-patch clamp combination, we would like to make the next big step in the integral understanding of our biological and synthetic systems in the context of three key projects. For each project, patch clamp measurements are carried out in parallel with various fluorescence microscopy techniques, e.g. Ca-imaging, FRET, or colocalization studies.For example, one project will shed light on the atypical activation of B cells by the bacterial lectins BambL from B. ambifaria and LecB from P. aeruginosa. This is induced by the binding of the lectins to glycosylated plasma membrane receptors, especially to the B cell receptor. With the help of the new equipment, we can characterize the whole of the processes in much more detail. The opening of Ca2+-channels in the plasma membrane (recorded by patch clamp) can thus be correlated with the binding to receptors, the uptake of lectins / bacteria (recorded by fluorescence) and the Ca2+-release into the cytosol of the cells (recorded by Ca-imaging).The purchase of this unique combination of devices would enable us to conduct our studies of biological infection processes at a scientifically excellent level using these correlative and complementary measurement methods.

我们申请荧光-膜片钳组合，主要用于风险组2的人类病原体的感染工作。这套设备在弗赖堡大学是独一无二的，对于Winfried Römer工作组未来的研究活动至关重要，以便能够继续进行顶级研究。Römer实验室探索细菌与各种哺乳动物细胞（如肺、皮肤和免疫细胞）相互作用的分子机制。在这里，重点是细菌凝集素，它识别宿主细胞质膜上的糖基化受体，并诱导不同的信号通路，影响细胞的生理-在积极和消极的意义。我们方法的优势在于，一方面，从复杂的细胞生物学出发，我们可以识别关键分子，有了这些知识，我们可以在更简单的合成系统中重建自然过程，从而更好地理解它们。许多重要的受体，如生长因子受体，细胞粘附分子，鞘糖脂和离子通道是这些细菌凝集素的靶点，这些凝集素对质膜的组织，动力学和稳定性有很大影响，我们必须认识到，单靠一种技术，如荧光显微镜，远远不足以充分描述细胞过程的复杂性。在荧光-膜片钳组合的帮助下，我们希望在三个关键项目的背景下，在对我们的生物和合成系统的整体理解方面迈出下一大步。对于每个项目，膜片钳测量与各种荧光显微镜技术平行进行，例如钙成像，FRET或共定位研究。例如，一个项目将阐明来自B的细菌凝集素BambL对B细胞的非典型激活。ambifaria和来自铜绿假单胞菌的LecB。这是由凝集素与糖基化质膜受体，特别是与B细胞受体的结合诱导的。在新设备的帮助下，我们可以更详细地描述整个过程。质膜Ca ~（2+）通道的开放（通过膜片钳记录）因此可以与受体的结合相关，凝集素/细菌的摄取（通过荧光记录）和Ca 2+释放到细胞的胞质溶胶中（通过钙成像记录）。购置这套独特的仪器组合，可使我们在科学上达到卓越的水平，研究生物感染过程使用这些相关和互补的测量方法。