Wright, Elizabeth R. and Iancu, Cristina V. and Tivol, William F. and Jensen, Grant J. (2006) Observations on the behavior of vitreous ice at ~82 and ~12K. Journal of Structural Biology, 153 (3). pp. 241-252. ISSN 1047-8477 http://resolver.caltech.edu/CaltechAUTHORS:20110803-130902686
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In an attempt to determine why cooling with liquid helium actually proved disadvantageous in our electron cryotomography experiments, further tests were performed to explore the differences in vitreous ice at ~82 and ~12 K. Electron diffraction patterns showed clearly that the vitreous ice of interest in biological electron cryomicroscopy (i.e., plunge-frozen, buffered protein solutions) does indeed collapse into a higher density phase when irradiated with as few as 2–3 e^−/Å^2 at ~12 K. The high density phase spontaneously expanded back to a state resembling the original, low density phase over a period of hours at ~82 K. Movements of gold fiducials and changes in the lengths of tunnels drilled through the ice confirmed these phase changes, and also revealed gross changes in the concavity of the ice layer spanning circular holes in the carbon support. Brief warmup–cooldown cycles from ~12 to ~82 K and back, as would be required by the flip-flop cryorotation stage, did not induce a global phase change, but did allow certain local strains to relax. Several observations including the rates of tunnel collapse and the production of beam footprints suggested that the high density phase flows more readily in response to irradiation. Finally, the patterns of bubbling were different at the two temperatures. It is concluded that the collapse of vitreous ice at ~12 K around macromolecules is too rapid to account alone for the problematic loss of contrast seen, which must instead be due to secondary effects such as changes in the mobility of radiolytic fragments and water.
|Additional Information:||Copyright © 2005 Elsevier Inc. Received 29 June 2005; received in revised form 16 November 2005; accepted 7 December 2005. Available online 5 January 2006. We thank H. Jane Ding for help preparing figures. This work was supported in part by NIH Grant PO1 GM66521 to G.J.J., DOE Grant DE-FG02-04ER63785 to G.J.J., a Searle Scholar Award to G.J.J., the Beckman Institute at Caltech, and gifts to Caltech from the Ralph M. Parsons Foundation, the Agouron Institute, and the Gordon and Betty Moore Foundation.|
|Subject Keywords:||Liquid helium; Radiation damage; Vitreous ice; Amorphous ice; Cryoelectron microscopy|
|Official Citation:||Elizabeth R. Wright, Cristina V. Iancu, William F. Tivol, Grant J. Jensen, Observations on the behavior of vitreous ice at ~82 and~12 K, Journal of Structural Biology, Volume 153, Issue 3, March 2006, Pages 241-252, ISSN 1047-8477, DOI: 10.1016/j.jsb.2005.12.003. (http://www.sciencedirect.com/science/article/pii/S1047847705002893)|
|Usage Policy:||No commercial reproduction, distribution, display or performance rights in this work are provided.|
|Deposited By:||George Porter|
|Deposited On:||03 Aug 2011 20:45|
|Last Modified:||03 Aug 2011 20:45|
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