01443nas a2200205 4500008004100000022001400041245010400055210006900159260001200228300001100240490000800251520078400259653002801043653004001071100001701111700002501128700001701153700001901170856004801189 2009 eng d a1095-865700aDetermination of signal-to-noise ratios and spectral SNRs in cryo-EM low-dose imaging of molecules.0 aDetermination of signaltonoise ratios and spectral SNRs in cryoE c05/2009 a126-320 v1663 a

Attempts to develop efficient classification approaches to the problem of heterogeneity in single-particle reconstruction of macromolecules require phantom data with realistic noise models. We have estimated the signal-to-noise ratios and spectral signal-to-noise ratios for three steps in the electron microscopic image formation from data obtained experimentally. An important result is that structural noise, i.e., the irreproducible component of the object prior to image formation, is substantial, and of the same order of magnitude as the reproducible signal. Based on this result, the noise modeling for testing new classification techniques can be improved.

10aCryoelectron Microscopy10aImage Processing, Computer-Assisted1 aBaxter, Bill1 aGrassucci, Robert, A1 aGao, Haixiao1 aFrank, Joachim uhttp://www.ncbi.nlm.nih.gov/pubmed/1926933201752nas a2200265 4500008004100000022001400041245010100055210006900156260001200225300001000237490000800247520093400255653002801189653002101217653002201238653004001260653001401300100002001314700002501334700002201359700001701381700002101398700001901419856004801438 2008 eng d a1095-865700aExploration of parameters in cryo-EM leading to an improved density map of the E. coli ribosome.0 aExploration of parameters in cryoEM leading to an improved densi c10/2008 a24-320 v1643 a

A number of image processing parameters in the 3D reconstruction of a ribosome complex from a cryo-EM data set were varied to test their effects on the final resolution. The parameters examined were pixel size, window size, and mode of Fourier amplitude enhancement at high spatial frequencies. In addition, the strategy of switching from large to small pixel size during angular refinement was explored. The relationship between resolution (in Fourier space) and the number of particles was observed to follow a lin-log dependence, a relationship that appears to hold for other data, as well. By optimizing the above parameters, and using a lin-log extrapolation to the full data set in the estimation of resolution from half-sets, we obtained a 3D map from 131,599 ribosome particles at 6.7A resolution (FSC=0.5).

10aCryoelectron Microscopy10aEscherichia coli10aImage Enhancement10aImage Processing, Computer-Assisted10aRibosomes1 aLeBarron, Jamie1 aGrassucci, Robert, A1 aShaikh, Tanvir, R1 aBaxter, Bill1 aSengupta, Jayati1 aFrank, Joachim uhttp://www.ncbi.nlm.nih.gov/pubmed/1860654902306nas a2200289 4500008004100000022001400041245013100055210006900186260001200255300000900267490000800276520139700284653001501681653002801696653001901724653002201743653004001765653002501805653002601830653001401856100002201870700002001892700002001912700001701932700001901949856004801968 2008 eng d a1095-865700aParticle-verification for single-particle, reference-based reconstruction using multivariate data analysis and classification.0 aParticleverification for singleparticle referencebased reconstru c10/2008 a41-80 v1643 a

As collection of electron microscopy data for single-particle reconstruction becomes more efficient, due to electronic image capture, one of the principal limiting steps in a reconstruction remains particle-verification, which is especially costly in terms of user input. Recently, some algorithms have been developed to window particles automatically, but the resulting particle sets typically need to be verified manually. Here we describe a procedure to speed up verification of windowed particles using multivariate data analysis and classification. In this procedure, the particle set is subjected to multi-reference alignment before the verification. The aligned particles are first binned according to orientation and are binned further by K-means classification. Rather than selection of particles individually, an entire class of particles can be selected, with an option to remove outliers. Since particles in the same class present the same view, distinction between good and bad images becomes more straightforward. We have also developed a graphical interface, written in Python/Tkinter, to facilitate this implementation of particle-verification. For the demonstration of the particle-verification scheme presented here, electron micrographs of ribosomes are used.

10aAlgorithms10aArtificial Intelligence10aClassification10aImage Enhancement10aImage Processing, Computer-Assisted10aMicroscopy, Electron10aMultivariate Analysis10aRibosomes1 aShaikh, Tanvir, R1 aTrujillo, Ramon1 aLeBarron, Jamie1 aBaxter, Bill1 aFrank, Joachim uhttp://www.ncbi.nlm.nih.gov/pubmed/1861954701670nas a2200289 4500008004100000022001400041245011900055210006900174260001200243300001200255490000600267520076600273653004001039653002501079653002201104653002401126653001301150653002801163100002201191700001701213700001701230700002701247700002101274700001801295700001901313856004801332 2008 eng d a1750-279900aSPIDER image processing for single-particle reconstruction of biological macromolecules from electron micrographs.0 aSPIDER image processing for singleparticle reconstruction of bio c10/2008 a1941-740 v33 a

This protocol describes the reconstruction of biological molecules from the electron micrographs of single particles. Computation here is performed using the image-processing software SPIDER and can be managed using a graphical user interface, termed the SPIDER Reconstruction Engine. Two approaches are described to obtain an initial reconstruction: random-conical tilt and common lines. Once an existing model is available, reference-based alignment can be used, a procedure that can be iterated. Also described is supervised classification, a method to look for homogeneous subsets when multiple known conformations of the molecule may coexist.

10aImage Processing, Computer-Assisted10aMicroscopy, Electron10aModels, Molecular10aMolecular Structure10aSoftware10aUser-Computer Interface1 aShaikh, Tanvir, R1 aGao, Haixiao1 aBaxter, Bill1 aAsturias, Francisco, J1 aBoisset, Nicolas1 aLeith, ArDean1 aFrank, Joachim uhttp://www.ncbi.nlm.nih.gov/pubmed/1918007801572nas a2200217 4500008004100000022001400041245004500055210004300100260001200143300001000155490000800165520098000173653002601153653004001179653001301219653002001232100001701252700001801269700001901287856004801306 2007 eng d a1047-847700aSPIRE: the SPIDER reconstruction engine.0 aSPIRE the SPIDER reconstruction engine c01/2007 a56-630 v1573 a

SPIRE is a Python program written to modernize the user interaction with SPIDER, the image processing system for electron microscopical reconstruction projects. SPIRE provides a graphical user interface (GUI) to SPIDER for executing batch files of SPIDER commands. It also lets users quickly view the status of a project by showing the last batch files that were run, as well as the data files that were generated. SPIRE handles the flexibility of the SPIDER programming environment through configuration files: XML-tagged documents that describe the batch files, directory trees, and presentation of the GUI for a given type of reconstruction project. It also provides the capability to connect to a laboratory database, for downloading parameters required by batch files at the start of a project, and uploading reconstruction results at the end of a project.

10aComputational Biology10aImage Processing, Computer-Assisted10aSoftware10aSoftware Design1 aBaxter, Bill1 aLeith, ArDean1 aFrank, Joachim uhttp://www.ncbi.nlm.nih.gov/pubmed/1705574302034nas a2200361 4500008004100000022001400041245010100055210006900156260001200225300001100237490000800248520093900256653001501195653002001210653002801230653005301258653004001311653003101351653001301382653001801395653001401413653002601427653001301453653002401466100001701490700001701507700002101524700001801545700001901563700001901582700002301601856004801624 2005 eng d a1047-847700aParticle picking by segmentation: a comparative study with SPIDER-based manual particle picking.0 aParticle picking by segmentation a comparative study with SPIDER c12/2005 a211-200 v1523 a

Boxing hundreds of thousands of particles in low-dose electron micrographs is one of the major bottle-necks in advancing toward achieving atomic resolution reconstructions of biological macromolecules. We have shown that a combination of pre-processing operations and segmentation can be used as an effective, automatic tool for identifying and boxing single-particle images. This paper provides a brief description of how this method has been applied to a large data set of micrographs of ice-embedded ribosomes, including a comparative analysis of the efficiency of the method. Some results on processing micrographs of tripeptidyl peptidase II particles are also shown. In both cases, we have achieved our goal of selecting at least 80% of the particles that an expert would select with less than 10% false positives.

10aAlgorithms10aAminopeptidases10aCryoelectron Microscopy10aDipeptidyl-Peptidases and Tripeptidyl-Peptidases10aImage Processing, Computer-Assisted10aImaging, Three-Dimensional10aInternet10aParticle Size10aRibosomes10aSerine Endopeptidases10aSoftware10aSoftware Validation1 aAdiga, Umesh1 aBaxter, Bill1 aHall, Richard, J1 aRockel, Beate1 aRath, Bimal, K1 aFrank, Joachim1 aGlaeser, Robert, M uhttp://www.ncbi.nlm.nih.gov/pubmed/1633022902292nas a2200301 4500008004100000022001400041245008900055210006900144260001200213300001100225490000800236520138600244653001501630653001501645653003001660653002801690653002201718653004001740653001801780653003501798653001401833653002001847100001701867700001801884700001701902700002301919856004801942 2004 eng d a1047-847700aA binary segmentation approach for boxing ribosome particles in cryo EM micrographs.0 abinary segmentation approach for boxing ribosome particles in cr c01/2004 a142-510 v1453 a

Three-dimensional reconstruction of ribosome particles from electron micrographs requires selection of many single-particle images. Roughly 100,000 particles are required to achieve approximately 10 A resolution. Manual selection of particles, by visual observation of the micrographs on a computer screen, is recognized as a bottleneck in automated single-particle reconstruction. This paper describes an efficient approach for automated boxing of ribosome particles in micrographs. Use of a fast, anisotropic non-linear reaction-diffusion method to pre-process micrographs and rank-leveling to enhance the contrast between particles and the background, followed by binary and morphological segmentation constitute the core of this technique. Modifying the shape of the particles to facilitate segmentation of individual particles within clusters and boxing the isolated particles is successfully attempted. Tests on a limited number of micrographs have shown that over 80% success is achieved in automatic particle picking.

10aAlgorithms10aAnisotropy10aAutomatic Data Processing10aCryoelectron Microscopy10aImage Enhancement10aImage Processing, Computer-Assisted10aParticle Size10aPattern Recognition, Automated10aRibosomes10aSoftware Design1 aAdiga, Umesh1 aMalladi, Ravi1 aBaxter, Bill1 aGlaeser, Robert, M uhttp://www.ncbi.nlm.nih.gov/pubmed/1506568105025nas a2200385 4500008004100000022001400041245011300055210006900168260001200237300001100249490000700260520391000267653002204177653001204199653002704211653002504238653002404263653001004287653002804297653004004325653002704365653002904392653001404421653001604435653001404451653001404465653001204479653002404491100001304515700001904528700001704547700001404564700001404578856004704592 1997 eng d a0735-109700aOptical mapping of drug-induced polymorphic arrhythmias and torsade de pointes in the isolated rabbit heart.0 aOptical mapping of druginduced polymorphic arrhythmias and torsa c03/1997 a831-420 v293 a

OBJECTIVES: 

This study sought to 1) test the hypothesis that in the setting of bradycardia and drug-induced action potential prolongation, multiple foci of early afterdepolarizations (EADs) result in beat to beat changes in the origin and direction of the excitation wave front and are responsible for polymorphic arrhythmias; and 2) determine whether EADs may initiate nonstationary reentry, giving rise to the typical torsade de pointes (TDP) pattern.

BACKGROUND: 

In the past, it has been difficult to associate EADs or reentry with the undulating electrocardiographic (ECG) patterns of TDP.

METHODS: 

A voltage-sensitive dye was used for high resolution video imaging of electrical waves on the epicardial and endocardial surface of the Langendorff-perfused rabbit heart. ECG and monophasic action potentials from the right septal region were also recorded. Bradycardia was induced by ablation of the atrioventricular node.

RESULTS: 

Perfusion of low potassium chloride Tyrode solution plus quinidine led to prolongation of the action potential and the QT interval. Eventually, EADs and triggered activity ensued, giving rise to intermittent episodes of polymorphic arrhythmia. In one experiment, triggered activity was followed by a long episode of vortex-like reentry with an ECG pattern characteristic of TDP. However, in most experiments, focal activity of varying origins and propagation patterns was observed. Triggered responses also showed varying degrees of local block. Similar results were obtained with E-4031. Burst pacing both at control conditions and in the presence of quinidine consistently led to vortex-like reentry whose ECG pattern resembled TDP. However, the cycle length of the arrhythmia with quinidine was longer than that for control ([mean +/- SEM] 194 +/- 12 vs. 132 +/- 8 ms, p < 0.03).

CONCLUSIONS: 

Drug-induced polymorphic ventricular arrhythmias may result from beat to beat changes in wave propagation patterns initiated by EADs or EAD-induced nonstationary reentrant activity. In contrast, burst pacing-induced polymorphic tachycardia in the presence or absence of drugs is the result of nonstationary reentrant activity.

10aAction Potentials10aAnimals10aAnti-Arrhythmia Agents10aArrhythmias, Cardiac10aElectrocardiography10aHeart10aHeart Conduction System10aImage Processing, Computer-Assisted10aModels, Cardiovascular10aOrgan Culture Techniques10aPerfusion10aPiperidines10aPyridines10aQuinidine10aRabbits10aTorsades de Pointes1 aAsano, Y1 aDavidenko, J M1 aBaxter, Bill1 aGray, R A1 aJalife, J uhttp://www.ncbi.nlm.nih.gov/pubmed/909153103382nas a2200349 4500008004100000022001400041245008900055210006900144260001200213300001100225490000700236520236800243653002202611653001502633653001202648653003502660653001602695653002402711653002602735653002102761653004002782653002702822653001002849653002502859653002002884100001702904700001902921700001402940700001702954700001402971856004702985 1997 eng d a0090-696400aTechnical features of a CCD video camera system to record cardiac fluorescence data.0 aTechnical features of a CCD video camera system to record cardia c07/1997 a713-250 v253 a

A charge-coupled device (CCD) camera was used to acquire movies of transmembrane activity from thin slices of sheep ventricular epicardial muscle stained with a voltage-sensitive dye. Compared with photodiodes, CCDs have high spatial resolution, but low temporal resolution. Spatial resolution in our system ranged from 0.04 to 0.14 mm/pixel; the acquisition rate was 60, 120, or 240 frames/sec. Propagating waves were readily visualized after subtraction of a background image. The optical signal had an amplitude of 1 to 6 gray levels, with signal-to-noise ratios between 1.5 and 4.4. Because CCD cameras integrate light over the frame interval, moving objects, including propagating waves, are blurred in the resulting movies. A computer model of such an integrating imaging system was developed to study the effects of blur, noise, filtering, and quantization on the ability to measure conduction velocity and action potential duration (APD). The model indicated that blurring, filtering, and quantization do not affect the ability to localize wave fronts in the optical data (i.e., no systematic error in determining spatial position), but noise does increase the uncertainty of the measurements. The model also showed that the low frame rates of the CCD camera introduced a systematic error in the calculation of APD: for cutoff levels > 50%, the APD was erroneously long. Both noise and quantization increased the uncertainty in the APD measurements. The optical measures of conduction velocity were not significantly different from those measured simultaneously with microelectrodes. Optical APDs, however, were longer than the electrically recorded APDs. This APD error could be reduced by using the 50% cutoff level and the fastest frame rate possible.

10aAction Potentials10aAlgorithms10aAnimals10aBody Surface Potential Mapping10aCalibration10aComputer Simulation10aElectric Conductivity10aFluorescent Dyes10aImage Processing, Computer-Assisted10aModels, Cardiovascular10aSheep10aVentricular Function10aVideo Recording1 aBaxter, Bill1 aDavidenko, J M1 aLoew, L M1 aWuskell, J P1 aJalife, J uhttp://www.ncbi.nlm.nih.gov/pubmed/923698304137nas a2200313 4500008004100000022001400041245013200055210006900187260001300256300001200269490000700281520321100288653001203499653002403511653001003535653004003545653002703585653001403612653001203626653002903638100001403667700001403681700001603695700001703711700001203728700001903740700001703759856004703776 1995 eng d a0009-732200aNonstationary vortexlike reentrant activity as a mechanism of polymorphic ventricular tachycardia in the isolated rabbit heart.0 aNonstationary vortexlike reentrant activity as a mechanism of po c05/1995  a2454-690 v913 a

BACKGROUND: 

Ventricular tachycardia may result from vortexlike reentrant excitation of the myocardium. Our general hypothesis is that in the structurally normal heart, these arrhythmias are the result of one or two nonstationary three-dimensional electrical scroll waves activating the heart muscle at very high frequencies.

METHODS AND RESULTS: 

We used a combination of high-resolution video imaging, electrocardiography, and image processing in the isolated rabbit heart, together with mathematical modeling. We characterized the dynamics of changes in transmembrane potential patterns on the epicardial surface of the ventricles using optical mapping. Image processing techniques were used to identify the surface manifestation of the reentrant organizing centers, and the location of these centers was used to determine the movement of the reentrant pathway. We also used numerical simulations incorporating Fitzhugh-Nagumo kinetics and realistic heart geometry to study how stationary and nonstationary scroll waves are manifest on the epicardial surface and in the simulated ECG. We present epicardial surface manifestations (reentrant spiral waves) and ECG patterns of nonstationary reentrant activity that are consistent with those generated by scroll waves established at the right and left ventricles. We identified the organizing centers of the reentrant circuits on the epicardial surface during polymorphic tachycardia, and these centers moved during the episodes. In addition, the arrhythmias that showed the greatest movement of the reentrant centers displayed the largest changes in QRS morphology. The numerical simulations showed that stationary scroll waves give rise to monomorphic ECG signals, but nonstationary meandering scroll waves give rise to undulating ECGs characteristic of torsade de pointes.

CONCLUSIONS: 

Polymorphic ventricular tachycardia in the healthy, isolated rabbit heart is the result of either a single or paired ("figure-of-eight") nonstationary scroll waves. The extent of the scroll wave movement corresponds to the degree of polymorphism in the ECG. These results are consistent with our numerical simulations that showed monomorphic ECG patterns of activity for stationary scroll waves but polymorphic patterns for scroll waves that were nonstationary.

10aAnimals10aElectrocardiography10aHeart10aImage Processing, Computer-Assisted10aModels, Cardiovascular10aPerfusion10aRabbits10aTachycardia, Ventricular1 aGray, R A1 aJalife, J1 aPanfilov, A1 aBaxter, Bill1 aCabo, C1 aDavidenko, J M1 aPertsov, A V uhttp://www.ncbi.nlm.nih.gov/pubmed/7729033