A new microscopy technique known as SUSHI (Super-resolution Shadow Imaging) has been designed to improve the imaging of cells in living brain tissue. The technique combines 3D-STED microscopy and fluorescent labeling of the extracellular fluid.
Microscopy is a basic tool in research into the biology of any organism given that the elements studied, the cells, are of microscopic and frequently nanoscopic size. Until now, existing microscopy methods to explore living brain tissue have been limited to imaging previously labelled cells only.
Yet, owing to technical limitations, not all the cells in a specific region of the brain can be labelled simultaneously; this has restricted the way we see and therefore understand how brain cells, which are highly interconnected, are organised and interact with each other.
Extracellular Fluid Labelling
SUSHI allows the extracellular fluid, the tiny space full of liquid surrounding brain cells to be labelled in one sweep, thus obviating the need to individually label all the cells that one is intending to analyse.
Because this “label” also remains outside the cells, a kind of negative image akin to the film used in old cameras is produced. So the negative image contains the same information about the brain cells as its corresponding positive image, but thanks to the fact that the labelling procedure is more straightforward, it is much easier to obtain this image and all the information contained in it.
The advance is the result of a cross-border, interdisciplinary project developed between the research group led by Professor Valentin Nägerl of the University of Bordeaux (France) and Dr Jan Tønnesen, UPV/EHU University of the Basque Country Department of Neurosciences. Dr. Tønnesen works at the ACHUCARRO facilities inside the university’s Science Park in Leioa.
“The SUSHI technique is revolutionary because it allows us to simultaneously image all the brain cells in a specific region of living brain tissue. In the past we used to come across blank spaces in the microscopy images, because we were unable to label all the cells at the same time. This fact was a big constraint for us. From now on, this technique will enable us to see all the cells in the area of study that we put under the microscope lens as well as all their interactions, and that will allow us to advance our knowledge of brain functions in a healthy organ and in a diseased one,”
said Dr Tønnesen.
Stimulated emission depletion (STED) microscopy is one of several types of super resolution microscopy techniques that have recently been developed to bypass the diffraction limit of light microscopy to increase resolution. STED is a deterministic functional technique that exploits the non-linear response of fluorophores commonly used to label biological samples in order to achieve an improvement in resolution, that is to say STED allows for images to be taken at resolutions below the diffraction limit.
The work was supported by grants from the Agence Nationale de la Recherche, France-BioImaging and the Foundation Recherche Medicale. Dr Tønnesen. was supported by an EMBO postdoctoral fellowship and a Ramon y Cajal fellowship.