Proteins under the microscope
A new device called the Volta phase plate could improve the quality of images taken using an electron microscope.
One way of investigating how proteins and other biological molecules work is to look at their structure. Light microscopes cannot produce detailed enough images to fully reveal these structures, and so a technique called cryo-electron microscopy is often used instead. In this technique, a biological sample is frozen to the temperature of liquid nitrogen and a beam of electrons is fired at it to create an image. By taking many of these images and then subjecting them to computer processing it is possible to reconstruct the three-dimensional structure of the molecule.
Frozen biological samples are essentially transparent to the electron beam used in an electron microscope. To view samples, researchers therefore use a method called phase contrast, which relies on a property of the electron beam (called its phase) changing as the beam passes through the sample. The traditional “defocus” method of producing phase contrast from electron microscopy relies on processing a series of slightly out-of-focus images of the sample.
Phase plates are add-on devices that are commonly used in light microscopes to produce phase contrast. For many years now, attempts have been made to produce a working phase plate for electron microscopes. However, an effective plate, called the Volta phase plate, has only recently been developed.
Danev and Baumeister have now evaluated how well the Volta phase plate performs during the analysis of a single, relatively large protein. This molecule is considered ‘easy’ to analyze using cryo-electron microscopy as relatively few microscopic images need to be recorded to solve the protein’s structure. Danev and Baumeister found that the Volta phase plate matched or slightly exceeded the performance of the traditional defocus method of producing phase contrast, depending on how many images were used to analyze the protein. This is the first time that a phase plate has matched the performance of the defocus method.
A future challenge will be to make the experimental procedures and the software involved in using the Volta phase plate more user-friendly. The phase plate also needs to be tested with more ‘difficult’ samples, such as small proteins and samples whose structure could not be established using the defocus method of producing phase contrast.
To find out more
Read the eLife research paper on which this eLife digest is based: “Cryo-EM single particle analysis with the Volta phase plate” (March 7, 2016).
Read a commentary on this research paper: “Cryo-EM: Protein complexes in focus”.
