Phase contrast electron microscopy: development of thin-film phase plates and biological applications.
Phase contrast transmission electron microscopy (TEM) based on thin-film phase plates has been developed and applied to biological systems. Currently, development is focused on two techniques that employ two different types of phase plates. The first technique uses a Zernike phase plate, which is made of a uniform amorphous carbon film that completely covers the aperture of an objective lens and can retard the phase of electron waves by pi/2, except at the centre where a tiny hole is drilled. The other technique uses a Hilbert phase plate, which is made of an amorphous carbon film that is twice as thick as the Zernike phase plate, covers only half of the aperture and retards the electron wave phase by pi. By combining the power of efficient phase contrast detection with the accurate preservation achieved by a cryotechnique such as vitrification, macromolecular complexes and supermolecular structures inside intact bacterial or eukaryotic cells may be visualized without staining. Phase contrast cryo-TEM has the potential to bridge the gap between cellular and molecular biology in terms of high-resolution visualization. Examples using proteins, viruses, cyanobacteria and somatic cells are provided.
Nagayama
Kuniaki
K
Okazaki Institute for Integrative Bioscience and National Institute for Physiological Sciences, National Institutes of Natural Sciences, Okazaki 444-8787, Japan. nagayama@nips.ac.jp
Danev
Radostin
R
D002477:Cells / Q000648:ultrastructure; D020285:Cryoelectron Microscopy / Q000379:methods*; D046529:Microscopy, Electron, Transmission / Q000295:instrumentation* / Q000379:methods*; D008858:Microscopy, Phase-Contrast / Q000295:instrumentation* / Q000379:methods*; D011506:Proteins / Q000648:ultrastructure; D014780:Viruses / Q000648:ultrastructure
D011506:Proteins