Tuning of the Zernike phase-plate for visualization of detailed ultrastructure in complex biological specimens Journal Article


Author(s): Fukuda, Yoshiyuki; Fukazawa, Yugo; Danev, Radostin S; Shigemoto, Ryuichi; Nagayama, Kuniaki
Article Title: Tuning of the Zernike phase-plate for visualization of detailed ultrastructure in complex biological specimens
Affiliation
Abstract: In order to acquire phase-contrast images with adequate contrast, conventional TEM requires large amount of defocus. Increasing the defocus improves the low-frequency components but attenuates the high-frequency ones. On the other hand, Zernike phase-contrast TEM (ZPC-TEM) can recover low-frequency components without losing the high-frequency ones under in-focus conditions. ZPC-TEM however, has another problem, especially in imaging of complex biological specimens such as cells and tissues; strong halos appear around specimen structures, and these halos hinder the interpretation of images. Due to this problem, the application of ZPC-TEM has been restricted to imaging of smaller particles. In order to improve the halo appearance, we fabricated a new quarter-wave thin film phase-plate with a smaller central hole and tested it on vitreous biological specimens. ZPC-TEM with the new plate could successfully visualize, in in-focus images, the intracellular fine features of cultured cells and brain tissues. This result indicates that reduction of the central hole diameter makes ZPC-TEM applicable on size scales ranging from protein particles to tissue sections. The application of ZPC-TEM to vitreous biological specimens will be a powerful method to advance the new field of imaging science for ultrastructures in close-to-physiological state.
Keywords: Brain tissue; Cryo-EM; Quarter-wave film phase-plate; Zernike phase-contrast TEM
Journal Title: Journal of Structural Biology
Volume: 168
Issue 3
ISSN: 1095-8657
Publisher: Academic Press  
Date Published: 2009-12-01
Start Page: 476
End Page: 484
DOI: 10.1016/j.jsb.2009.08.011
Open access: no