scholarly journals Using the Volta phase plate with defocus for cryo-EM single particle analysis

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Radostin Danev ◽  
Dimitry Tegunov ◽  
Wolfgang Baumeister
Keyword(s):  
Data Acquisition ◽  
Single Particle ◽  
Hybrid Approach ◽  
Particle Analysis ◽  
Phase Plate ◽  
Single Particle Analysis ◽  
Resolution Limit ◽  
Contrast Transfer Function ◽  
Acquisition Speed ◽  
Acquisition Method

Previously, we reported an in-focus data acquisition method for cryo-EM single-particle analysis with the Volta phase plate (Danev and Baumeister, 2016). Here, we extend the technique to include a small amount of defocus which enables contrast transfer function measurement and correction. This hybrid approach simplifies the experiment and increases the data acquisition speed. It also removes the resolution limit inherent to the in-focus method thus allowing 3D reconstructions with resolutions better than 3 Å.

scholarly journals Using the Volta phase plate with defocus for cryo-EM single particle analysis

2016 ◽  
Author(s):  
Radostin Danev ◽  
Dimitry Tegunov ◽  
Wolfgang Baumeister
Keyword(s):  
Data Acquisition ◽  
Single Particle ◽  
Hybrid Approach ◽  
Particle Analysis ◽  
Phase Plate ◽  
Single Particle Analysis ◽  
Function Measurement ◽  
Contrast Transfer Function ◽  
Acquisition Speed ◽  
Acquisition Method

Previously, we reported an in-focus data acquisition method for cryo-EM single particle analysis with the Volta phase plate (VPP) (Danev and Baumeister, 2016). Here, we extend the technique to include a small amount of defocus which enables contrast transfer function measurement and correction. This hybrid approach simplifies the experiment and increases the data acquisition speed. It also enables 3D reconstructions with resolutions in the 2 A range, demonstrating that, in practice, there are no resolution limitations imposed by the VPP.

scholarly journals Advances in Single Particle Analysis Data Acquisition

2019 ◽  
Vol 25 (S2) ◽  
pp. 1012-1013 ◽  
Author(s):  
Stan Konings ◽  
Maarten Kuijper ◽  
Jeroen Keizer ◽  
Fanis Grollios ◽  
Tjerk Spanjer ◽  
...  
Keyword(s):  
Data Acquisition ◽  
Single Particle ◽  
Analysis Data ◽  
Particle Analysis ◽  
Single Particle Analysis

scholarly journals Zernike Phase Plate Cryoelectron Microscopy Facilitates Single Particle Analysis of Unstained Asymmetric Protein Complexes

Structure ◽  
2010 ◽  
Vol 18 (1) ◽  
pp. 17-27 ◽  
Author(s):  
Wei-Hau Chang ◽  
Michael T.-K. Chiu ◽  
Chin-Yu Chen ◽  
Chi-Fu Yen ◽  
Yen-Cheng Lin ◽  
...  
Keyword(s):  
Single Particle ◽  
Protein Complexes ◽  
Cryoelectron Microscopy ◽  
Particle Analysis ◽  
Phase Plate ◽  
Single Particle Analysis

A3Near-atomic Resolution Single Particle Analysis with the Volta Phase Plate

Microscopy ◽  
2015 ◽  
Vol 64 (suppl 1) ◽  
pp. i9.2-i9
Author(s):  
Radostin Danev ◽  
Maryam Khoshouei ◽  
Wolfgang Baumeister
Keyword(s):  
Single Particle ◽  
Atomic Resolution ◽  
Particle Analysis ◽  
Phase Plate ◽  
Single Particle Analysis ◽  
Resolution Single

scholarly journals Phase Contrast Cryo-Electron Tomography and Single Particle Analysis with a New Phase Plate

2014 ◽  
Vol 20 (S3) ◽  
pp. 232-233
Author(s):  
Maryam Khoshouei ◽  
Radostin Danev ◽  
Günther Gerisch ◽  
Maria Ecke ◽  
Juergen Plitzko ◽  
...  
Keyword(s):  
Single Particle ◽  
Phase Contrast ◽  
Electron Tomography ◽  
Particle Analysis ◽  
Phase Plate ◽  
Single Particle Analysis ◽  
Cryo Electron Tomography ◽  
New Phase

scholarly journals Automated Cryo-tomography and Single Particle Analysis with a New Type of Phase Plate

2014 ◽  
Vol 20 (S3) ◽  
pp. 206-207
Author(s):  
Radostin Danev ◽  
Bart Buijsse ◽  
Yoshiyuki Fukuda ◽  
Maryam Khoshouei ◽  
Juergen Plitzko ◽  
...  
Keyword(s):  
Single Particle ◽  
Particle Analysis ◽  
Phase Plate ◽  
Single Particle Analysis ◽  
New Type

scholarly journals Cryo-EM single particle analysis with the Volta phase plate

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Radostin Danev ◽  
Wolfgang Baumeister
Keyword(s):  
Single Particle ◽  
Spherical Aberration ◽  
Quality Data ◽  
Particle Analysis ◽  
Limiting Factor ◽  
Phase Plate ◽  
Single Particle Analysis ◽  
High Quality Data ◽  
Size Heterogeneity ◽  
Resolution Single

We present a method for in-focus data acquisition with a phase plate that enables near-atomic resolution single particle reconstructions. Accurate focusing is the determining factor for obtaining high quality data. A double-area focusing strategy was implemented in order to achieve the required precision. With this approach we obtained a 3.2 Å resolution reconstruction of the Thermoplasma acidophilum 20S proteasome. The phase plate matches or slightly exceeds the performance of the conventional defocus approach. Spherical aberration becomes a limiting factor for achieving resolutions below 3 Å with in-focus phase plate images. The phase plate could enable single particle analysis of challenging samples in terms of small size, heterogeneity and flexibility that are difficult to solve by the conventional defocus approach.

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