Journal of Structural Biology, Vol.151, No.3, 288-297, 2005
A "flip-flop" rotation stage for routine dual-axis electron cryotomography
Electron cryotomography can be used to solve the three-dimensional structures of individual large macromolecules, assemblies, and even small intact cells to medium (similar to 4-8 nm) resolution in a near-native state, but restrictions in the range of accessible views are a major limitation. Here we report on the design, characterization, and demonstration of a new "flip-flop" rotation stage that allows facile and routine collection of two orthogonal tilt-series of cryosamples. single- and dual-axis tomograrns of a variety of samples are compared to illustrate qualitatively the improvement produced by inclusion of the second tilt-series. Exact quantitative expressions are derived for the volume of the remaining "missing pyramid" in reciprocal space. When orthogonal tilt-series are recorded to +/-65 degrees in each direction, as this new cryostage permits, only 11% of reciprocal space is left unmeasured. The tomograms suggest that further improvement could be realized, however, through better software to align and merge dual-axis tilt-series of cryosamples. (C) 2005 Elsevier Inc. All rights reserved.