Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

A test-bed for optimizing high-resolution single particle reconstructions.

Scott M Stagg1, Gabriel C Lander, Joel Quispe

  • 1The National Resource for Automated Molecular Microscopy, Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.

Journal of Structural Biology
|June 7, 2008
PubMed
Summary
This summary is machine-generated.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

<i>Siphoviridae</i> phage tails co-enrich with ex vivo amyloids.

bioRxiv : the preprint server for biology·2026
Same author

Helical reconstruction of amyloids in cryoSPARC.

Acta crystallographica. Section F, Structural biology communications·2026
Same author

Characterization and modulation of human insulin degrading enzyme conformational dynamics to control enzyme activity.

eLife·2026
Same author

Characterization Standard for <i>In-situ</i> Cryo-electron Tomography.

bioRxiv : the preprint server for biology·2026
Same author

AreTomoLive: automated reconstruction of comprehensively corrected and denoised cryo-electron tomograms in real time and at high throughput.

Nature methods·2026
Same author

Three-Dimensional Visualization and Proteomic Analysis of Human Cardiac Transthyretin Amyloidosis Tissue Reveals Microangiopathy and Capillary Occlusion.

Journal of the American Heart Association·2026
Same journal

Ultrastructural evidence of autophagy-related processes and mitochondrial remodeling in the myxozoan parasite Henneguya piaractus.

Journal of structural biology·2026
Same journal

Architecture and dynamics of a supramolecular oxygen transport system in human homogentisate 1,2-Dioxygenase.

Journal of structural biology·2026
Same journal

Connecting pathways between mineralized fibrocartilage and bone at the Achilles tendon insertion.

Journal of structural biology·2026
Same journal

Structural and functional characterization of thermostable EstS1 esterase for BHET degradation.

Journal of structural biology·2026
Same journal

Following the white rabbit: multiscale 2D3D correlative imaging of bone structure.

Journal of structural biology·2026
Same journal

The mantis shrimp eye imaged in 3D using 4th generation synchrotron multiscale phase contrast tomography.

Journal of structural biology·2026
See all related articles

Optimizing cryo-electron microscopy (cryo-EM) parameters significantly improves 3D reconstruction resolution. This study achieved a 5.4A map, revealing secondary structures and alpha-helix handedness in GroEL.

Area of Science:

  • Structural Biology
  • Biophysics
  • Microscopy

Background:

  • Cryo-electron microscopy (cryo-EM) single particle reconstruction routinely achieves ~10Å resolution, with <5Å being exceptional.
  • Practical resolution limits in cryo-EM are often attributed to specimen preparation, data collection, and analysis, not microscope resolving power (<2Å).

Purpose of the Study:

  • To systematically optimize parameters for high-resolution single particle reconstruction in cryo-EM.
  • To identify key factors influencing resolution in 3D electron density map generation.

Main Methods:

  • Employed a highly automated image acquisition system and relational database for parameter tracking.
  • Performed systematic 3D reconstructions of GroEL, varying particle count, accelerating voltage, and electron dose.

Related Experiment Videos

  • Investigated methods for excluding "bad" particles from final map computation.

    • Main Results:

    • Achieved a 5.4Å nominal resolution (FSC(0.5)) cryo-EM map of GroEL using standard instrumentation and an automated approach.
    • The resulting map clearly discerns secondary structures.
    • The handedness of some alpha-helices within the GroEL structure was determined.

    Conclusions:

    • Systematic optimization of cryo-EM parameters, including automated data processing and particle selection, can significantly enhance 3D reconstruction resolution.
    • Achieving sub-6Å resolution is feasible with standard equipment, enabling detailed structural analysis like secondary structure identification.