Analysis and sorting by flow cytometry

Hugo Berthelot, a postdoc on the Pepperkok team, is studying the diverse shapes of these diazotrophs and other cyanobacteria, such as Liqueria intracellularis, which coexist with Hemiaurus. Flow cytometry allows him to classify and study different cells.Credits: Sebastien Colin / Max Planck Institute for Developmental Biology, Tübingen

Everywhere in the ocean of the planet, there are small groups of creatures doing fairly monumental work.

Known as diazotrophs, these special bacteria convert atmospheric nitrogen into biologically usable forms that are essential for all living organisms to live. As the ocean warms and the pH balance changes, scientists want to know more about these little creatures that make a significant contribution to our survival as species. Often, the first step in these investigations is flow Sight meter.

That is certainly the case with Hugo Berthelot, a postdoc of the Peppelcock team studying the various forms of diazotrophs. He is interested in whether their shape affects how well nitrogen-fixing bacteria convert nitrogen or the ability of habitats to withstand climate change.

Berthelot is also one of more and more EMBL researchers whose work at the EMBL flow cytometry facility is more than just analyzing and classifying human cells to reflect the expanding One Health perspective.

“We are just beginning to understand the diversity of these cells,” Berthelot said. He uses a flow cytometer to classify cells before performing genomic and other omics analyzes. For example, better understand where in the cell photosynthesis takes place.

“Liquidity” of flow cytometer

Cytometers and cell sorters use laser beams to analyze, classify, and separate cells or organisms in a stable fluid stream.

Diana Ordonez, Head of Flow Cytometry Core Facility at EMBL Heidelberg, said:

“This device produces droplets containing particles or cells that are excited by a laser beam, measures the emitted light, and then deflects the droplets that carry the particles of interest from the stream to a collection tube or plate. “She explained. “It makes it possible to separate a given particle.”

For example, scientists can isolate a population of interest, such as T cells, from a blood sample that contains several types of immune cells and use it for downstream analysis.

Ordonez and Gerald Pfister, heads of the flow cytometry facility at the EMBL Roman Epigenetics and Neurobiology Unit, agree that if a sample can be suspended in a liquid, it can be analyzed with a flow cytometer.

“The most typical example is when doing a blood test. Cell populations in the blood are identified and quantified using a flow cytometer,” says Pfister. “But the flow cytometer is a food for assessing algae cells, cilia (protozoa with hairy edges known as cilia), bacteria in water samples, as well as bacteria in milk and yeast cells in beer. It is used to look up various things, such as quality control systems.

Flow cytometry and bridge construction

Ordonez, who has been working at the EMBL Heidelberg Flow Cytometry Facility since 2015, took command in February.

Pfister arrived at the EMBL Rome facility in June after playing various roles in industry and academia around the world, including Austria, Brazil, Belgium and Qatar, for over 20 years. In Qatar, he led the development of the Flow Cytometer Core Lab. In all his roles, he has paid attention to end-user services and public support.

“My hope is to be actively involved in the project here,” Fister said. “At this point, I have a lot of tips and ideas, so people can really contribute when planning a project.”

Ordonez also has a long history of flow cytometry. Before coming to EMBL, she used flow cytometry at the Center d’immunologie de Marseille-Luminy to use a variety of mouse immunological cells (T cells, B cells, dendritic cells, natural killer cells, monospheres, progenitor cells). I studied cells). Her goal was to establish a reliable model for analyzing the role that certain types of white blood cells play in the immune response.

After that, she moved to EMBL. The main reason is that, like Pfister, he enjoys the training, advice and support of other researchers in flow cytometry. It is this attitude that guides both scientists in sharing their plans for the future.

Future outlook

As EMBL embarks on its next five-year research program, EMBL’s flow cytometry facility is looking for ways to adapt to an increasingly interdisciplinary environment and will begin exploring biological systems in natural conditions. ..

“I have a PhD in freshwater biology and am looking forward to this challenge,” said Pfister. “This opens up opportunities to collaborate with a variety of research teams, as well as those related to biomedicine and medical conditions.”

The two EMBL facilities have flow cytometers and cell sorters with slightly different capabilities. Ordonez and Pfister perform most of the flow cytometry work themselves, advise individual researchers on specific projects, develop optimal flow applications, and troubleshoot experimental setups.

Training helps expand the pool of hands-on users and allows others to learn how flow cytometry can be used in their various research projects. From data analysis workshops to hands-on courses in flow cytometry and cell sorting presented in collaboration with the German Cancer Research Center (DFKZ), EMBL’s flow cytometry training is available to the international scientific community and is always available. There is a high demand. That’s why Ordonez is looking for creative ways to extend it.

With teamwork, flow cytometry works … better

Collaboration with other EMBL core facilities has been part of that creative management approach in the past and may play a role in the future. Ordonez quickly finds a convenient location in the Genomics Core Facility (GeneCore), directly opposite the hall for gene sequencing. This is a logical follow-up to flow cytometry for epigenetics and genetics researchers.

Over the years, the challenging scientific needs of EMBL’s flow cytometry customers have made these facilities excellent testbeds for piloting new technologies. In fact, Berthelot’s research benefits from “beta testing” of just this kind of new equipment. He uses detectors to create rapid-fire images and uses them in addition to traditional flow cytometric parameters to analyze and separate particles of interest. These images help Berthelot more easily identify and understand the complexity of the organisms present in the sample.

Ordonez and Pfister agree that the two facilities will work together to find more and more ways to enhance their offerings and improve their position at EMBL. “I had this dream of repairing an oil spill, so I started studying microbiology,” Ordonez said. “I ended up studying immunology, but we all need to do something for this planet. It just has to be done. Here with all these resources of the EMBL A bright heart can make a difference. ”

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Quote: Https: // Analysis and sorting by flow cytometry (October 22, 2021) obtained on October 22, 2021

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