Spot-scan imaging of ice-embedded catalase crystal on a slow-scan CCD camera in a 400-kV electron cryomicroscope

1994 ◽  
Vol 52 ◽  
pp. 98-99
Author(s):  
Wah Chiu ◽  
Michael Sherman ◽  
Jaap Brink
Keyword(s):  
Electron Diffraction ◽  
Ccd Camera ◽  
Protein Crystal ◽  
Modulation Transfer ◽  
Measured Intensity ◽  
3 Dimensional ◽  
Dimensional Reconstruction ◽  
Diffraction Patterns ◽  
Complex Crystal

In protein electron crystallography, both low dose electron diffraction patterns and images are needed to provide accurate amplitudes and phases respectively for a 3-dimensional reconstruction. We have demonstrated that the Gatan 1024x1024 model 679 slow-scan CCD camera is useful to record electron diffraction intensities of glucose-embedded crotoxin complex crystal to 3 Å resolution. The quality of the electron diffraction intensities is high on the basis of the measured intensity equivalence ofthe Friedel-related reflections. Moreover, the number of patterns recorded from a single crystal can be as high as 120 under the constraints of radiation damage and electron statistics for the reflections in each pattern.A limitation of the slow-scan CCD camera for recording electron images of protein crystal arises from the relatively large pixel size, i.e. 24 μm (provided by Gatan). The modulation transfer function of our camera with a P43 scintillator has been determined for 400 keV electrons and shows an amplitude fall-off to 0.25 at 1/60 μm−1.

Merging of electron-diffraction intensities acquired with a slow-scan CCD camera of crotoxin complex crystals to 3.5Å resolution

1994 ◽  
Vol 52 ◽  
pp. 104-105
Author(s):  
Jaap Brink ◽  
Wah Chiu
Keyword(s):  
Electron Diffraction ◽  
Ccd Camera ◽  
Crystal Thickness ◽  
Local Contrast ◽  
Camera Model ◽  
Data Set ◽  
3 Dimensional ◽  
Diffraction Mode ◽  
Diffraction Patterns ◽  
Complex Crystals

Crotoxin complex is the principal neurotoxin of the South American rattlesnake, Crotalus durissus terrificus and has a molecular weight of 24 kDa. The protein is a heterodimer with subunit A assigneda chaperone function. Subunit B carries the lethal activity, which is exerted on both sides ofthe neuro-muscular junction, and which is thought to involve binding to the acetylcholine receptor. Insight in crotoxin complex’ mode of action can be gained from a 3 Å resolution structure obtained by electron crystallography. This abstract communicates our progress in merging the electron diffraction amplitudes into a 3-dimensional (3D) intensity data set close to completion. Since the thickness of crotoxin complex crystals varies from one crystal to the other, we chose to collect tilt series of electron diffraction patterns after determining their thickness. Furthermore, by making use of the symmetry present in these tilt data, intensities collected only from similar crystals will be merged.Suitable crystals of glucose-embedded crotoxin complex were searched for in the defocussed diffraction mode with the goniometer tilted to 55° of higher in a JEOL4000 electron cryo-microscopc operated at 400 kV with the crystals kept at -120°C in a Gatan 626 cryo-holder. The crystal thickness was measured using the local contrast of the crystal relative to the supporting film from search-mode images acquired using a 1024 x 1024 slow-scan CCD camera (model 679, Gatan Inc.).

Anisotropic radiation damage of potassium tetracyano platinate (KCP)

1978 ◽  
Vol 36 (1) ◽  
pp. 392-393
Author(s):  
N. Uyeda ◽  
E. J. Kirkland ◽  
B. M. Siegel
Keyword(s):  
Electron Diffraction ◽  
Radiation Damage ◽  
Structural Changes ◽  
High Resolution Electron Microscopy ◽  
Radiation Sensitivity ◽  
Resolution Electron ◽  
Damage Process ◽  
Diffraction Patterns ◽  
Fading Rate

The direct observation of structural change by high resolution electron microscopy will be essential for the better understanding of the damage process and its mechanism. However, this approach still involves some difficulty in quantitative interpretation mostly being due to the quality of obtained images. Electron diffraction, using crystalline specimens, has been the method most frequently applied to obtain a comparison of radiation sensitivity of various materials on the quantitative base. If a series of single crystal patterns are obtained the fading rate of reflections during the damage process give good comparative measures. The electron diffraction patterns also render useful information concerning the structural changes in the crystal. In the present work, the radiation damage of potassium tetracyano-platinate was dealt with on the basis two dimensional observation of fading rates of diffraction spots. KCP is known as an ionic crystal which possesses “one dimensional” electronic properties and it would be of great interest to know if radiation damage proceeds in a strongly asymmetric manner.

Modulation Transfer Function Measurement System at Different Contrasts for CCD Camera

2012 ◽  
Vol 241-244 ◽  
pp. 648-652
Author(s):  
Wen Juan Li ◽  
Chuan Liang ◽  
Ning Zhi Jin ◽  
Mei Lan Zhou
Keyword(s):  
Measurement System ◽  
Ccd Camera ◽  
Imaging Systems ◽  
Integrating Sphere ◽  
Test Target ◽  
Modulation Transfer ◽  
Imaging Quality ◽  
The Face ◽  
The Many

In order to achieve MTFs at different contrasts, the MTF measurement system is designed and developed. Two integrating spheres are used to illuminate the face and back of the test target uniformly. The target luminance and background luminance of the test target are regulated by adjusting the attenuators near the entrance of each integrating sphere. The many groups’ experimental results indicate that the luminance differences between the values by the system and those by L88 first level luminance meter, which is checked by National Institute of Metrology P. R. China, are within ±0.3 cd/m2. Thereby the measurement precision can be ensured in MTF test. MTFs of Sony camera and Cannon camera at different contrasts are measured by this system. The measurement values show that MTFs at different contrasts can demonstrate the imaging quality fully and objectively. This study provides an effective method to evaluate the imaging quality of visible imaging systems.

Automation in electron diffraction analysis

1995 ◽  
Vol 53 ◽  
pp. 170-171
Author(s):  
J. M. Zuo
Keyword(s):  
Electron Diffraction ◽  
Diffraction Analysis ◽  
Diffraction Pattern ◽  
Ccd Camera ◽  
Lattice Parameter ◽  
Electron Diffraction Analysis ◽  
Full Automation ◽  
Imaging Plates ◽  
Complex Crystal

Automated lattice parameter measurement and orientation analysis are often needed in the characterization of microstructures using electron diffraction, and is made possible with increasingly popular use of slow scan CCD camera and imaging plates. Both of these two detectors are largely linear and digital. Typical electron diffraction analysis has three steps: 1) diffraction pattern measurement, 2) diffraction pattern indexing and 3) solution. Full automation in all these three steps is desired, however, may be hard to achieve especially for complex crystal structures. The importance of automation in each of these steps depends on the type of analysis and the number of analysis needed. Full automation is necessary in the type of applications where the same analysis is repeated many times, such as in texture analysis. In table 1, various applications of electron diffraction and automation needed are listed.There are two types of approach to the automatic indexing. The commonly used method is to matching a calculated list of g's with the measured ones.

Electron-diffraction patterns from frozen hydrate protein microcrystals: A new tool instructural studies

1994 ◽  
Vol 52 ◽  
pp. 102-103
Author(s):  
Christoph Burmester ◽  
Kenneth C. Holmes ◽  
Rasmus R. Schröder
Keyword(s):  
Electron Diffraction ◽  
Diffraction Data ◽  
Heavy Atom ◽  
Atomic Resolution ◽  
Protein Crystals ◽  
Electron Diffraction Data ◽  
Phase Information ◽  
Isomorphous Replacement ◽  
Diffraction Patterns

Electron crystallography of 2D protein crystals can yield models with atomic resolution by taking Fourier amplitudes from electron diffraction and phase information from processed images. Imaging at atomic resolution is more difficult than the recording of corresponding electron diffraction patterns. Therefore attempts have been made to recover phase information from diffraction data from 2-D and 3-D crystals by the method of isomorphous replacement using heavy atom labelled protein crystals. These experiments, however, have so far not produced usable phase information, partly because of the large experimental error in the spot intensities. Here we present electron diffraction data obtained from frozen hydrated 3-D protein crystals with an energy-filter microscope and a specially constructed Image Plate scanner which are of considerably better crystallographic quality (as evidenced in much smaller values for the crystallographic R-factors Rsym and Rmerge) than those reported before. The quality of this data shows that the method of isomorphous replacement could indeed be used for phase determination for diffraction data obtained from 3-D microcrystals by electron diffraction.

Electron Microscopy and Electron Diffraction of Hydrated Biological Material

1974 ◽  
Vol 32 ◽  
pp. 296-297
Author(s):  
S.W. Hui ◽  
D.F. Parsons
Keyword(s):  
Electron Microscopy ◽  
Phase Transition ◽  
Fine Structure ◽  
Electron Diffraction ◽  
Biological Material ◽  
Physiological State ◽  
Phospholipid Bilayers ◽  
Protein Crystal ◽  
Surface Polysaccharides ◽  
Diffraction Patterns

The importance of electron microscopy and electron diffraction of hydrated biological material needs no stress. The diffraction patterns of wet and stained catalase obtained at this laboratory shows that the fine structure of the protein crystal is destroyed by the usual drying and staining processes. The different phase transition behaviors of wet and dry phospholipid bilayers, as studied by electron diffraction, illustrates the necessity of studying the biological material in a more physiological state. Cell surface polysaccharides can be studied by electron microscopy only if they are preserved in a wet environment. An ultimate achievement of the hydration stage work would be visualization of living processes in an electron microscope.

Electron cryo-microscopy of macromolecular assembly at 400 kV

1991 ◽  
Vol 49 ◽  
pp. 426-427
Author(s):  
Wah Chiu ◽  
Michael F. Schmid ◽  
Joanita Jakana ◽  
Paul Matsudaira
Keyword(s):  
Crystallographic Analysis ◽  
Biological Macromolecules ◽  
Electron Microscopic ◽  
Low Contrast ◽  
3 Dimensional ◽  
Macromolecular Assembly ◽  
High Resolution Structure ◽  
Dimensional Reconstruction ◽  
Computational Procedures

Electron cryo-microscopy has proven to be a valuable technique for determining 3-dimensional structures of biological macromolecules. The cryo technique is capable of preserving biological specimens in their native conformation and of reducing radiation damage during the microscopic observation. Computer processing is used to combine data from different angular views for 3-dimensional reconstruction. The structural detail revealed in this type of analysis can be limited by the quality of the electron microscopic images and the computational procedures used to retrieve the low contrast signals. So far, the highest resolution study of electron cryo-microscopy is bacteriorhodopsin where the polypeptide chain can be traced and some of the amino acid side chains can be identified. Though high resolution structure can be obtained by electron crystallographic analysis, further improvement can be made in several areas. These include specimen flatness, specimen movement induced by the electrons, and achievement of better signal to noise ratios in the images.

Quantitative Electron Diffraction for Crystal Structure Determination

2009 ◽  
Vol 1184 ◽  
Author(s):  
Peter Oleynikov ◽  
Daniel Grüner ◽  
Daliang Zhang ◽  
Junliang Sun ◽  
Xiaodong Zou ◽  
...  
Keyword(s):  
Electron Diffraction ◽  
Data Quality ◽  
Diffraction Data ◽  
Ccd Camera ◽  
Electron Diffraction Data ◽  
X Ray Diffraction ◽  
Quantitative Investigation ◽  
Critical Question ◽  
Selected Area Electron Diffraction

AbstractWe present a quantitative investigation of data quality using electron precession, compared to standard selected-area electron diffraction (SAED). Data can be collected on a CCD camera and automatically extracted by computer. The critical question of data quality is addressed – can electron diffraction data compete with X-ray diffraction data in terms of resolution, completeness and quality of intensities?

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