AsianScientist (Jun. 16, 2017) – A team of researchers at the Okinawa Institute of Science and Technology Graduate University (OIST) have developed a one-of-a-kind microscope that uses graphene to enhance microscopic images of minuscule viruses. Their findings have been published in Microscopy.
Electron microscopes rely on an electron beam rather than light to illuminate the target sample at a much higher resolution. The electron beam hits the sample, allowing scientists to build an accurate image of the target based on the scattering of electrons.
While conventional electron microscopes require high-energy electrons, the microscope described in the present study focuses on low-energy electrons which can potentially be much more efficient at imaging viruses if the associated technical issues can be overcome.
“Low-energy electrons interact very strongly with matter,” explained Dr. Masao Yamashita, the first author of the study. “They are great for imaging biological specimens, made up of light materials like carbon, oxygen and nitrogen, which are basically transparent to high energy electrons.”
Using low-energy electrons however has an important drawback: because of its high sensitivity with matter, a low-energy electron beam would interact with the target sample but also with everything else like the support plate and film on which the sample is laying. The resulting image would not distinguish the study material from the background.
To counter this effect, the researchers from the Quantum Wave Microscopy Unit turned to the unique properties of graphene. They synthesized a film made of a single layer—one atom thin—of graphene on which the biological samples, like the viruses they study, will be displayed.
Graphene is extremely conductive, which means electrons can cross the layer very easily. This way, the low energy electrons will interact very little with the background graphene layer and much more with the virus sample which will stand out with a great contrast. This high conductivity also prevents ‘charging-up,’ an accumulation of electrons on the film that would distort the final image. The thinness of the film also provides a much brighter background—thus a much better contrast with the study material—than conventional carbon films.
“The graphene film allows us to achieve great contrast with very low energy electrons, allowing to enhance tiny details,” added Yamashita.
However, graphene film is not so easy to handle. It is so clear that it needs to be spotless and free from any contaminants, leading the OIST scientists to develop a technique to meticulously clean the graphene film.
There is also an issue with loading the virus sample onto the graphene film. The graphene film is oily, while biological preparations would typically be in water-based. They would not mix very well: if you just add the viruses onto the film, the results are viruses clinging together in scattered dense spots making it impossible to reveal individual details.
To solve this second issue, the researchers resorted to using centrifugal force to scatter the viruses onto the entire surface of the film, preventing them from making clumps. The viruses are loaded in a tube with the graphene film sitting at one end, while the other end is attached to a vertical axis which is spun up to 100,000 rotations per minute. The centrifugal force pushes the viruses onto the graphene film and prevents them from regrouping, allowing the researchers to see distinctive details on each specimen.
The outcome of all these efforts is higher resolution pictures of viruses’ shells, whose shape and morphological details can provide clues on how to fight them. Using graphene and a low-voltage electron beam, the researchers reveal minuscule details of bacteriophage T4 that were previously invisible on conventional carbon film, like the fiber-like limbs that the virus uses to hook onto its bacterial prey.
Yamashita and his team are already working on the next step to further increase the quality of the images. To reconstruct images and to study the morphology of various types of specimen in the future, the ability to confidently compare microscopic images of biological materials at such a tiny scale require very high consistency between samples. To achieve this condition, the researchers are now developing a robust way to prepare the viruses by spraying them onto the graphene film in a sterile vacuum.
The article can be found at: Yamashita et al. (2017) Graphene Specimen Support Technique for Low Voltage STEM Imaging.
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Source: Okinawa Institute of Science and Technology Graduate University.
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