5-laser flow cytometer

• Subject Area: Neurosciences
• Term: Funded in 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 453253304

A key method in immunology and more recently also neurobiology is multicolour flow cytometry, which aims at detailed phenotyping and functional characterization of various cell populations. Due to the complexity of relevant immune cell subsets such as T lymphocytes, dendritic cells or monocytes/macrophages, this requires high-parameter measurements to dissect underlying disease mechanisms. Since patient material for such analyses is in general limited, combining 20+ parameters within a single measurement is extremely important. Along these lines, analysing up to 30 parameters simultaneously will help to reduce the number of animals that are required to tackle a specific research question. Both for human and animal studies, access to a state-of-the-art flow cytometer will allow a comprehensive analysis of even rare immune cell subsets as measurements will not be limited to identification of these cells from various tissues and body fluids, but will give additional information on functional properties such as cytokine secretion or activation status. Further, novel transgenic mouse models, as well as reporter system for cell culture assays rely on the expression of red fluorescent proteins. Currently, these reporter systems cannot be used as they require excitation by a yellow-green laser, which is not available in our LSR II flow cytometer. Replacing the LSR II device by a novel 5-laser-system will allow to combine a huge variety of fluorophores targeting surface markers as well as cytoplasmic and nuclear proteins with fluorochrome-based reporter systems and probes for detection of dead cells or RNA hybridisation. Access to five laser lines will allow us to spread our markers of interest over a wide spectrum, which is crucial to minimize spill-over effects and hence reduce compensation. Thereby, the sensitivity to detect weak signals is massively increased. The replacement of our current LSR II device by a modern high-parameter flow cytometer is key to the endeavour to dissect the underlying pathomechanisms of various neuroinflammatory and neurodegenerative diseases, which will ultimately help to develop novel therapeutic strategies.

• DFG Programme: Major Research Instrumentation
• Major Instrumentation: 5-Laser Durchflusszytometer
• Instrumentation Group: 3500 Zellzähl- und Klassiergeräte (außer Blutanalyse), Koloniezähler
• Applicant Institution: Universität Hamburg
• Leader: Professor Dr. Manuel A. Friese