The Electron Microscopy eXchange (EMX) initiative.

The cryo-EM structure of ETB-Gi complex, obtained using the Fusion-G system, provides insights into the distinct activation mechanism and G-protein promiscuity of ETB, aiding in the design of ETB agonists for therapeutic applications.

BackgroundThe transmission electron microscope is used to acquire structural information of macromolecular complexes. However, as any other imaging device, it introduces optical aberrations that must be corrected if high-resolution structural information is to be obtained. The set of all aberrations are usually modeled in Fourier space by the so-called Contrast Transfer Function (CTF). Before correcting for the CTF, we must first estimate it from the electron micrographs. This is usually done by estimating a number of parameters specifying a theoretical model of the CTF. This estimation is performed by minimizing some error measure between the theoretical Power Spectrum Density (PSD) and the experimentally observed PSD. The high noise present in the micrographs, the possible local minima of the error function for estimating the CTF parameters, and the cross-talking between CTF parameters may cause errors in the estimated CTF parameters.ResultsIn this paper, we explore the effect of these estimation errors on the theoretical CTF. For the CTF model proposed in [1] we show which are the most sensitive CTF parameters as well as the most sensitive background parameters. Moreover, we provide a methodology to reveal the internal structure of the CTF model (which parameters influence in which parameters) and to estimate the accuracy of each model parameter. Finally, we explore the effect of the variability in the detection of the CTF for CTF phase and amplitude correction.ConclusionWe show that the estimation errors for the CTF detection methodology proposed in [1] does not show a significant deterioration of the CTF correction capabilities of subsequent algorithms. All together, the methodology described in this paper constitutes a powerful tool for the quantitative analysis of CTF models that can be applied to other models different from the one analyzed here.

ACE: Automated CTF Estimation

Image processing in three-dimensional electron microscopy (3D-EM) is characterized by large amounts of data, and voluminous computing requirements. Here, we report our first experience with grid computing in this area. We present an interface between grid computing middleware and our image processing package Xmipp. The efficacy of this approach was illustrated with an Xmipp application for estimation of the contrast transfer function. In addition, we report our experience with grid computing in the development of a novel image refinement algorithm based on maximum likelihood principles. Its extensive CPU-requirements might have seriously hampered the algorithm development, if not for the far-reaching resources of grid computing. Our results suggest that electron microscopy image processing may be particularly well suited for grid computing.

Chromatic and achromatic (AC) assessments of camouflage textiles have been critical to the defense researchers for concealment, detection, recognition, and identification (CDRI) of target signature against multidimensional combat background (CB). AC assessment and camouflage measurement techniques are simulated and experimented for assessment of camouflage textiles against CB. This model has been demonstrated for color measurement spectrophotometer, scanning electron microscopy (SEM), digital imaging, hyperspectral imaging, and image processing software (ImageJ) for the advancement and establishment of AC camouflage textiles assessment. The chromatic variations of 48 artificial target objects (TOBs) have been synthesized by image processing; the technique can be implemented for defense CB-CDRI assessment. Microstructural variation versus optical signal of woodland, desertland and stoneland CB materials have been elucidated by SEM magnification. The achromatic variation of CB materials have been demonstrated for the replacement of optical signal against modern remote sensing device to the imaging sensor. Color difference (Δ E), microstructural variations, pixel variations to imaging signal and standard deviation of CB materials have been represented for remote sensing surveillance of defense applications against TOB-CB-CDRI. Technical simulation of color, texture, gloss, and pixel intensity has been derived for AC-CDRI assessment of camouflage textiles in TOBs-CB environment.

Image formation in phase-contrast electron microscopy is governed by the contrast transfer function (CTF). The key parameter to tune the CTF is the defocus. A precise and unbiased estimate of the defocus is essential to determine the forward model and interpret high resolution images in cryo-TEM. We present an algorithm based on the weak-phase approximation for determination of the defocus and astigmatism of the objective lens from recorded images of an amorphous sample. The algorithm identifies Thon rings in the power spectrum density (PSD) and uses them to estimate astigmatism and defocus together with their uncertainties. For the astigmatism estimation we use a transformation to polar coordinates to fit ellipses to the Thon rings. Averaging the PSD over these ellipses reduces CTF estimation to a 1D problem and enhances the signal-to-noise ratio.

Purpose The dicentric chromosome assay (DCA), recognized as the gold standard for biological dosimetry in emergency radiation exposure, involves multiple key steps. Among these, metaphase image selection remains ambiguous and particularly challenging for beginners. This study evaluates the impact of metaphase image selection on dicentric (Dic) frequency during initial training. Materials and methods Peripheral blood (PB) from three healthy donors was irradiated with 2 Gy X-rays and incubated at 37 °C for 2 h for DNA repair. PB samples were then cultured for 48 h with phytohemagglutinin and colcemid. After fixation, metaphase spreads were prepared, Giemsa-stained, and images were captured by specific microscope imaging and image processing software. Three beginners independently selected and analyzed 50 metaphase images each. A trainer with experience in performing radiation emergency related DCA then reviewed and removed unsuitable images. The beginners reanalyzed Dic frequencies, supplementing with additional images as needed to reach 50 metaphases. Dic frequencies were compared before and after image refinement. Results Dic frequencies were lower in metaphase image pools selected by beginners compared to those refined by the trainer. The novices’ selections included numerous over-condensed metaphases, making Dic detection difficult. Conclusion Blood culture for 48 h permanent colcemid treatment increases the occurrence of over-condensed chromosomes, affecting Dic scoring. This study highlights the importance of proper training in metaphase image selection and structured education on quality assessment, including the use of specific microscope imaging and image processing software to ensure accurate biological dosimetry.

Correction of axial chromatic aberration improves the contrast transfer in bright-field phase contrast imaging

Abstract: Contrast transfer function (CTF) is a vital function in transmission electron microscopy (TEM). It expresses to what extent amplitudes converted from the phase changes of the diffracted waves contribute to the TEM image, including the effects of lens aberrations. Simulation is very helpful to understand the application of the function thoroughly. In this work, we develop the CTFscope as a component in the Landyne software suite, to calculate the CTF with temporal and spatial dumping envelopes for conventional TEM and to extend it to various aberrations (up to fifth order) for aberration-corrected (AC)-TEM. It also includes effects on the CTF and imaging due to the objective aperture and image drift for tutorial purposes. The CTFscope has a user-friendly graphical interface for simulation, visualization, saving, and loading of the microscopy conditions and results. It can be used to explore instrumental performance information due to optical aberrations, effects on the resolution of a TEM and AC-TEM, and for teaching electron microscopy courses.

Application of template matching technique to particle detection in electron micrographs

Quantifying resolution limiting factors in subtomogram averaged cryo-electron tomography using simulations.

Structure projection retrieval by image processing of HREM images taken under non-optimum defocus conditions

The theoretical outcomes and experimental results of new color model implemented in algorithms and software of image processing are presented in the paper. This model, as it will be shown below, may be used in modern real time video processing applications such as radar tracking and communication systems. The developed model allows carrying out the image process with the least time delays (i.e. it speeding up image processing). The proposed model can be used to solve the problem of true color object identification Experimental results show that the time spent during RGI color model conversion may approximately four times less than the time spent during other similar models. .

The pyruvate dehydrogenase multienzyme complexes are among the largest multifunctional catalytic machines in cells, catalyzing the production of acetyl CoA from pyruvate. We have previously reported the molecular architecture of an 11-MDa subcomplex comprising the 60-mer icosahedral dihydrolipoyl acetyltransferase (E2) decorated with 60 copies of the heterotetrameric (alpha(2)beta(2)) 153-kDa pyruvate decarboxylase (E1) from Bacillus stearothermophilus (Milne, J. L. S., Shi, D., Rosenthal, P. B., Sunshine, J. S., Domingo, G. J., Wu, X., Brooks, B. R., Perham, R. N., Henderson, R., and Subramaniam, S. (2002) EMBO J. 21, 5587-5598). An annular gap of approximately 90 A separates the acetyltransferase catalytic domains of the E2 from an outer shell formed of E1 tetramers. Using cryoelectron microscopy, we present here a three-dimensional reconstruction of the E2 core decorated with 60 copies of the homodimeric 100-kDa dihydrolipoyl dehydrogenase (E3). The E2E3 complex has a similar annular gap of approximately 75 A between the inner icosahedral assembly of acetyltransferase domains and the outer shell of E3 homodimers. Automated fitting of the E3 coordinates into the map suggests excellent correspondence between the density of the outer shell map and the positions of the two best fitting orientations of E3. As in the case of E1 in the E1E2 complex, the central 2-fold axis of the E3 homodimer is roughly oriented along the periphery of the shell, making the active sites of the enzyme accessible from the annular gap between the E2 core and the outer shell. The similarities in architecture of the E1E2 and E2E3 complexes indicate fundamental similarities in the mechanism of active site coupling involved in the two key stages requiring motion of the swinging lipoyl domain across the annular gap, namely the synthesis of acetyl CoA and regeneration of the dithiolane ring of the lipoyl domain.

We present MOPEX - a software package for astronomical image processing and display. The package is a combination of command-line driven image processing software written in C/C++ with a Java-based GUI. The main image processing capabilities include creating mosaic images, image registration, background matching, point source extraction, as well as a number of minor image processing tasks. The combination of the image processing and display capabilities allows for much more intuitive and efficient way of performing image processing. The GUI allows for the control over the image processing and display to be closely intertwined. Parameter setting, validation, and specific processing options are entered by the user through a set of intuitive dialog boxes. Visualization feeds back into further processing by providing a prompt feedback of the processing results. The GUI also allows for further analysis by accessing and displaying data from existing image and catalog servers using a virtual observatory approach. Even though originally designed for the Spitzer Space Telescope mission, a lot of functionalities are of general usefulness and can be used for working with existing astronomical data and for new missions. The software used in the package has undergone intensive testing and benefited greatly from effective software reuse. The visualization part has been used for observation planning for both the Spitzer and Herschel Space Telescopes as part the tool Spot. The visualization capabilities of Spot have been enhanced and integrated with the image processing functionality of the command-line driven MOPEX. The image processing software is used in the Spitzer automated pipeline processing, which has been in operation for nearly 3 years. The image processing capabilities have also been tested in off-line processing by numerous astronomers at various institutions around the world. The package is multi-platform and includes automatic update capabilities. The software package has been developed by a small group of software developers and scientists at the Spitzer Science Center. It is available for distribution at the Spitzer Science Center web page.