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J Struct Biol. 2007 Jan;157(1):148-55. Epub 2006 Nov 3.

Automated acquisition of electron microscopic random conical tilt sets.

Author information

1
The Howard Hughes Medical Institute, University of California, San Francisco, CA 94158-2517, USA.

Abstract

Single particle reconstruction using the random conical tilt data collection geometry is a robust method for the initial determination of macromolecular structures by electron microscopy. Unfortunately, the broad adoption of this powerful approach has been limited by the practical challenges inherent in manual data collection of the required pairs of matching high and low tilt images (typically 60 degrees and 0 degrees). The microscopist is obliged to keep the imaging area centered during tilting as well as to maintain accurate focus in the tilted image while minimizing the overall electron dose, a challenging and time consuming process. To help solve these problems, we have developed an automated system for the rapid acquisition of accurately aligned and focused tilt pairs. The system has been designed to minimize the dose incurred during alignment and focusing, making it useful in both negative stain and cryo-electron microscopy. The system includes a feature for montaging untilted images to ensure that all of the particles in the tilted image may be used in the reconstruction.

PMID:
17169745
PMCID:
PMC2556511
DOI:
10.1016/j.jsb.2006.10.026
[Indexed for MEDLINE]
Free PMC Article

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