Cryogenic x-ray diffraction microscopy utilizing high-pressure cryopreservation.
We present cryo x-ray diffraction microscopy of high-pressure-cryofixed bacteria and report high-convergence imaging with multiple image reconstructions. Hydrated D. radiodurans cells were cryofixed at 200 MPa pressure into ∼10-μm-thick water layers and their unstained, hydrated cellular environments were imaged by phasing diffraction patterns, reaching sub-30-nm resolutions with hard x-rays. Comparisons were made with conventional ambient-pressure-cryofixed samples, with respect to both coherent small-angle x-ray scattering and the image reconstruction. The results show a correlation between the level of background ice signal and phasing convergence, suggesting that phasing difficulties with frozen-hydrated specimens may be caused by high-background ice scattering.
Photon Sciences, Brookhaven National Laboratory, Upton, NY, 11973 USA.
Enju
E
Lima
European Synchrotron Radiation Facility, 71, avenue des Martyrs 38000 Grenoble, France.
Yuriy
Y
Chushkin
European Synchrotron Radiation Facility, 71, avenue des Martyrs 38000 Grenoble, France.
Peter
P
van der Linden
Department of Physics, Ulsan National Institute of Science and Technology, Ulsan 689-798, Republic of Korea and Cornell High Energy Synchrotron Source, Ithaca, NY 14853 USA.
Chae Un
CU
Kim
European Synchrotron Radiation Facility, 71, avenue des Martyrs 38000 Grenoble, France.
Federico
F
Zontone
European Synchrotron Radiation Facility, 71, avenue des Martyrs 38000 Grenoble, France.
Philippe
P
Carpentier
Cornell High Energy Synchrotron Source, Ithaca, NY 14853 USA and Department of Physics, Cornell University, Ithaca, NY 14853 USA.
Sol M
SM
Gruner
European Synchrotron Radiation Facility, 71, avenue des Martyrs 38000 Grenoble, France.
Petra
P
Pernot
D015925:Cryopreservation / Q000379:methods*; D034301:Deinococcus; D007053:Ice; D007091:Image Processing, Computer-Assisted; D008853:Microscopy / Q000379:methods*; D011312:Pressure; D053838:Scattering, Small Angle; D014961:X-Ray Diffraction / Q000379:methods*; D014965:X-Rays
D007053:Ice