Microscopy and Microanalysis

Biological Imaging: Techniques Development and Applications

Cryomesh™: A New Substrate for Cryo-Electron Microscopy

Craig Yoshiokaa1, Bridget Carraghera1 and Clinton S. Pottera1 c1

a1 The National Resource for Automated Molecular Microscopy, The Scripps Research Institute, La Jolla, CA 92037, USA

Abstract

Here we evaluate a new grid substrate developed by ProtoChips Inc. (Raleigh, NC) for cryo-transmission electron microscopy. The new grids are fabricated from doped silicon carbide using processes adapted from the semiconductor industry. A major motivating purpose in the development of these grids was to increase the low-temperature conductivity of the substrate, a characteristic that is thought to affect the appearance of beam-induced movement (BIM) in transmission electron microscope (TEM) images of biological specimens. BIM degrades the quality of data and is especially severe when frozen biological specimens are tilted in the microscope. Our results show that this new substrate does indeed have a significant impact on reducing the appearance and severity of beam-induced movement in TEM images of tilted cryo-preserved samples. Furthermore, while we have not been able to ascertain the exact causes underlying the BIM phenomenon, we have evidence that the rigidity and flatness of these grids may play a major role in its reduction. This improvement in the reliability of imaging at tilt has a significant impact on using data collection methods such as random conical tilt or orthogonal tilt reconstruction with cryo-preserved samples. Reduction in BIM also has the potential for improving the resolution of three-dimensional cryo-reconstructions in general.

(Received October 05 2009)

(Accepted November 30 2009)

Correspondence:

c1 Corresponding author. E-mail: cpotter@scripps.edu