Welcome to the new version of CaltechAUTHORS. Login is currently restricted to library staff. If you notice any issues, please email coda@library.caltech.edu
Published July 2014 | Supplemental Material + Accepted Version
Journal Article Open

Correlated cryogenic photoactivated localization microscopy and cryo-electron tomography

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
Files (65.1 MB)
Name Size Download all
md5:c1beff3a34f0573315f526db796010c6
26.7 MB Download
md5:57300a75e1dd78eec82746fe0bedb68d
865.2 kB Preview Download
md5:d0cbb838263678989efe47da967ccb91
11.0 MB Preview Download
md5:b6f3513d00ddc5035f73f645e3ba9732
26.5 MB Download

Additional details

Created:
August 20, 2023
Modified:
October 26, 2023