Confocal Laser Scanning Microscope for vertical imaging

• Subject Area: Plant Sciences
• Term: Funded in 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 499026372

Confocal microscopy-based live cell imaging enables dedicated time-lapse imaging of thicker specimens and is hence a powerful tool to address biological function through the study of cellular dynamics. Available microscopes largely limit live cell imaging applications in plant science, because they are either “upright” or “inverted” systems and thus the specimens are always examined horizontally. As a result, plant specimens are not placed along their actual growth axis and, hence, may experience a vectorial change in gravity, which induces alterations in hormone distribution along plant organs. Specifically, the detailed assessment of tropistic growth is hardly possible under these conditions. This limitation also restricts live cell imaging of gravity sensitive lateral rooting as well as nodule development. On the other hand, unintentional gravitropic stimuli limits the detailed live cell imaging of other processes, including growth towards light, as well as root zonation/differentiation and overall growth behaviour. A custom-made, by 90° tilted confocal microscope and dedicated tracking software would alleviate these limitations. This set up would empower our plant science community, enabling us to study plant specimens at high resolution at their vertical growth direction, thereby substantially reducing unwanted side effects of the imaging approach. We also envision to establish a tilted fluorescence stereomicroscope. Using this system, in vitro grown seedlings can be imaged here at much higher throughput when compared to the confocal system, allowing to pre-screen the appropriate samples and/or conditions. This also adds additional value, because specimens can be also imaged in vertically grown, soil-filled rhizotrons. This system will close a currently existing technical gap, enabling cell biological approaches in soil grown roots. To further advance sample throughput, we will develop additional methods that allows the system to automatically detect root tips. This will automate the imaging of multiple roots and its three-dimensional (3D) depiction. In addition, the obtained data will be stored, processed and quantified simultaneously on a server (remote processing server). This provides a high degree of data security and enables simultaneous data quantification. The platform is technically feasib and the LIC (Life Imaging Center) has the necessary expertise to implement this project. The platform for vertical imaging and ist potential for larger set ups will generate additional value for the overall research environment in Freiburg. Besides, it will support research on root system architecture, which is an important focus of the CIBSS Cluster of Excellence.

• DFG Programme: Major Research Instrumentation
• Major Instrumentation: Confokales Laser Scanning Mikroskop für vertikales Imaging
• Instrumentation Group: 5090 Spezialmikroskope
• Applicant Institution: Albert-Ludwigs-Universität Freiburg
• Leader: Professor Dr. Jürgen Kleine-Vehn