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A new fluorescence microscopy technique has produced the world’s first nanoscale 3D images of molecules in a whole, living cell, researchers at KTH Royal Institute of Technology reported.
Ilaria Testa, an associate professor at KTH and researcher at the Science for Life Laboratory, says the technique is capable of generating images with precision that until now has been exclusive to electron microscopy. The progress was reported today in Nature Biotechnology, with near-molecular scale images of proteins in the brain’s hippocampal neurons.
Dubbed 3D pRESOLFT, the KTH technique can envision proteins on a wider scale than possible with electron microscopy; and it may do so without killing the cell and sectioning it – two necessary steps with an electron microscope.
This technique allows us to image proteins with a completely new level of 3D spatial details, and importantly in situ within the cell.”
Ilaria Testa, Associate Professor, Researcher, Science for Life Laboratory, KTH The Royal Institute of Technology
“Here, we don’t have to do that. The cells are happily moving and undergoing important functions,” Ilaria says.
Ilaria’s laboratory developed the method as part of its focus on exploring the localization and function of neuronal proteins, especially in synapses and axons, where trafficking organelles and protein complexes are so crowded that they need to be visualized in high resolution in order to be examined.
In traditional fluorescence microscopy, visible light is used to illuminate cells and tissues that are colored with a fluorescent dye – but the method is limited to creating 2D images, typically at low resolution. 3D pRESOLFT improves upon the method using a combination of interference patterns between switchable fluorescence dye which can be turned off and on, like a light switch, though a large volume of parallel images are recorded. The sample is subjected to less mild complete, preventing the sample from fading.
The ability to look at living cells in 3D with such precision makes it feasible to study how proteins get involved in important and poorly known physiological processes, ” she says.
“We now can see the 3D structure of brain cells, and examine molecules that we consider important for learning and memory formation – and understand how they change location and shape when subjected to specific stimulations,” she says.
KTH The Royal Institute of Technology
Bodén, A., et al. (2021) Volumetric live cell imaging with three-dimensional parallelized RESOLFT microscopy. Nature Biotechnology. doi.org/10.1038/s41587-020-00779-2.