Published July 2014
| Supplemental Material + Accepted Version
Journal Article
Open
Correlated cryogenic photoactivated localization microscopy and cryo-electron tomography
Chicago
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Abstract
Cryo-electron tomography (CET) produces three-dimensional images of cells in a near-native state at macromolecular resolution, but identifying structures of interest can be challenging. Here we describe a correlated cryo-PALM (photoactivated localization microscopy)-CET method for localizing objects within cryo-tomograms to beyond the diffraction limit of the light microscope. Using cryo-PALM-CET, we identified multiple and new conformations of the dynamic type VI secretion system in the crowded interior of Myxococcus xanthus.
Additional Information
© 2014 Macmillan Publishers. Received 06 February 2014;Accepted 10 April 2014; Published online 11 May 2014. We thank A.W. McDowall, C. Oikonomou, A. Konovalova, L. Cai and T. Zhiyentayev for assistance and discussions. This work was supported in part by US National Institutes of Health grant R01 GM094800B to G.J.J., the Howard Hughes Medical Institute and the Max Planck Society. Contributions: Y.-W.C. and G.J.J. conceived the cryo-PALM idea. Y.-W.C. and S.C. configured the optical system. Y.-W.C., S.C. and E.I.T. tested fluorophores for photoactivatability at low temperatures. Y.-W.C. improved stability of cryo-FLM stage, prepared samples, overcame laser-induced ice crystallization on the sample, acquired and analyzed cryo-PALM data and conducted correlated cryo-PALM-CET. A.T.-L., S.L. and L.S.-A. generated M. xanthus strains and conducted functional analyses. Y.-W.C. and G.J.J. wrote the paper with input from all authors. Competing financial interests: Y.-W.C., S.C., E.I.T. and G.J.J. are affiliated with the California Institute of Technology, which has filed a patent application based on this work.Attached Files
Accepted Version - nihms587569.pdf
Supplemental Material - nmeth.2961-S1.pdf
Supplemental Material - nmeth.2961-sv1.mov
Supplemental Material - nmeth.2961-sv2.mov
Files
nihms587569.pdf
Additional details
- PMCID
- PMC4081473
- Eprint ID
- 44615
- Resolver ID
- CaltechAUTHORS:20140402-152401360
- NIH
- R01 GM094800B
- Howard Hughes Medical Institute (HHMI)
- Max Planck Society
- Created
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2014-06-25Created from EPrint's datestamp field
- Updated
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2021-11-10Created from EPrint's last_modified field