Innovations in microscopy have deepened our understanding of complex biological processes. Indeed, the pioneering work of Santiago Ramón y Cajal observing the microstructures of neurons gave rise to the field of neuroscience. More recent technological breakthroughs hold the potential to further expand our knowledge of neuronal networks and the internal structures of neurons. However, one limitation of all the currently available microscopy methods is that they do not allow the 3-D visualization of the cellular structures in the brain. The ability to view large networks of neurons and analyze their connections in microscopic detail has the potential to open up new research possibilities to better understand brain function. Now, using a newly developed method by
researchers at MIT, biological samples can be “inflated” prior to imaging, allowing the user to observe neuronal processes in three dimensions.
Expansion Microscopy (ExM) is a new method of magnifying cells and tissues, to a high resolution, with standard microscopy. Fixed mouse brain tissue was embedded with the building components for a polyacrylate gel. Polyacrylate gels consist of charged cross-linked polymer networks immersed in a fluid. One unique property of these gels is that when soaked in an aqueous solution (i.e. water), these gels can expand uniformly.
After the brains had been evenly infused with the gel, antibodies with fluorescent labels were constructed that bind both to the polymer matrix and the tissue. The brains were digested away leaving the fluorescent antibodies in place. The matrix was then submerged in water causing the resultant polymer network to expand. Impressively, Chen et al. were able to expand the brains about 5 times the normal size while maintaining the general morphology. They could visualize cultured neurons at a resolution of about 70 nm. ExM compares favorably with super-resolution microscopy techniques, which enable resolution within a range of 20-70 nm. However, one advantage of ExM over super-resolution microscopy is that ExM can be performed with a standard microscope and thus data can be generated much more quickly.
With ExM, scientists now have access to a new level of magnification via enhancement of biological samples. This high-resolution three-dimensional microscopy facilitates the structural analysis of neuronal structures in a readily accessible manner. What is exciting about this creative microscopy technique is the different ways that ExM can be applied to understand such profound neuronal
networks better. Although a standard microscope was used to achieve high-resolution magnification, the polymer network produced via ExM is also an ideal candidate for light sheet microscopy. With light sheet microscopy one can use optical sectioning and illuminate an object from the side, with a thin sheet of light. The enlarged brain reveals the intricate network of dendrites, neurons and
synapses allowing scientists to put together a three-dimensional model of the brain. Future fine-tuning of this technique could also permit the expansion of the brain to more than 10 times. In more ways than one, ExM expands our ability to probe the molecular keystones of physiological function and disease.
Sources:
Fei Chen, Paul W. Tillberg and Edward S. Boyden. Optical Imaging. Science. 2015. 347(6221):543-8.
Hans-Ulrich Dodt. The superresolved brain. Science. 2015. 347(6221):474-5.
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