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ABSTRACT: Electron tomography (ET) has been proven an essential technique for imaging the structure of cells beyond the range of the light microscope down to the molecular level. Large-field high-resolution views of biological specimens span more than four orders of magnitude in spatial scale, and, as a consequence, are rather difficult to generate directly. Various techniques have been developed towards generating those views, from increasing the sensor array size to implementing serial sectioning and montaging. Datasets and reconstructions obtained by the latter techniques generate multiple three-dimensional (3D) reconstructions, that need to be combined together to provide all the multiscale information. In this work, we show how to implement montages within TxBR, a tomographic reconstruction software package. This work involves some new application of mathematical concepts related to volume preserving transformations and issues of gauge ambiguity, which are essential problems arising from the nature of the observation in an electron microscope. The purpose of TxBR is to handle those issues as generally as possible in order to correct for most distortions in the 3D reconstructions and allow for a seamless recombination of ET montages.
Journal of Structural Biology 06/2012; 180(1):154-64. · 3.41 Impact Factor
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ABSTRACT: Data acquisition of cryo-electron tomography (CET) samples described in previous chapters involves relatively imprecise mechanical motions: the tilt series has shifts, rotations, and several other distortions between projections. Alignment is the procedure of correcting for these effects in each image and requires the estimation of a projection model that describes how points from the sample in three-dimensions are projected to generate two-dimensional images. This estimation is enabled by finding corresponding common features between images. This chapter reviews several software packages that perform alignment and reconstruction tasks completely automatically (or with minimal user intervention) in two main scenarios: using gold fiducial markers as high contrast features or using relevant biological structures present in the image (marker-free). In particular, we emphasize the key decision points in the process that users should focus on in order to obtain high-resolution reconstructions.
Methods in enzymology 01/2010; 482:343-67. · 1.90 Impact Factor
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CSIE 2009, 2009 WRI World Congress on Computer Science and Information Engineering, March 31 - April 2, 2009, Los Angeles, California, USA, 7 Volumes; 01/2009
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CSIE 2009, 2009 WRI World Congress on Computer Science and Information Engineering, March 31 - April 2, 2009, Los Angeles, California, USA, 7 Volumes; 01/2009
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ABSTRACT: Electron microscope tomography [1] has been proven as an essential technique for imaging the structure of cells beyond the range of the light microscope down to the molecular level. However, because of the extreme difference in spatial scales, there is a large gap to be bridged between light and electron microscopy. Various techniques have been developed, including increasing size of the sensor arrays, serial sectioning and montaging. Data sets and reconstructions obtained by the latter techniques generate many 3D reconstructions that need to be glued together to provide information at a larger spatial scale. However, during the course of data acquisition, thin slices may become warped in optical and electron microscope preparations. We review some procedures for de-warping sections and reassembling them into larger reconstructions, and present some data from electron microscopy.
Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference 01/2009; 2009:5772-6.
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ABSTRACT: Image alignment is a critical step in obtaining high quality reconstructions. Bundle adjustment, based on a general projective model, combined with calculation of the envelope of backprojected tangents to a surface of revolution around the rotation axis establish the parameters of the projection maps, rotation angles and axis of rotation. Subsequent regression techniques may be used to calculate higher order polynomial corrections to the projection maps, and can compensate for curvilinear trajectories through the object, sample warping, and optical aberration. Backprojection from properly filtered images along the nominal electron trajectories yields high quality 3D reconstructions. We report here on recent extensions to the electron microscope tomography code, TxBR. In addition we discuss the basis for this code in the theory of Fourier integral operators.
Image and Signal Processing, 2008. CISP '08. Congress on; 06/2008
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Methods in cell biology 02/2007; 79:643-60. · 2.05 Impact Factor
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ABSTRACT: Alignment of the individual images of a tilt series is a critical step in obtaining high-quality electron microscope reconstructions. We report on general methods for producing good alignments, and utilizing the alignment data in subsequent reconstruction steps. Our alignment techniques utilize bundle adjustment. Bundle adjustment is the simultaneous calculation of the position of distinguished markers in the object space and the transforms of these markers to their positions in the observed images, along the bundle of particle trajectories along which the object is projected to each EM image. Bundle adjustment techniques are general enough to encompass the computation of linear, projective or nonlinear transforms for backprojection, and can compensate for curvilinear trajectories through the object, sample warping, and optical aberration. We will also report on new reconstruction codes and describe our results using these codes.
Journal of Structural Biology 06/2006; 154(2):144-67. · 3.41 Impact Factor
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ABSTRACT: We describe the technique and application of energy filtering, automated most-probable loss (MPL) tomography to intermediate voltage electron microscopy (IVEM). We show that for thick, selectively stained biological specimens, this method produces a dramatic increase in resolution of the projections and the computed volumes versus standard unfiltered transmission electron microscopy (TEM) methods. This improvement in resolution is attributed to the reduction of chromatic aberration, which results from the large percentage of inelastic electron-scattering events for thick specimens. These improvements are particularly evident at the large tilt angles required to improve tomographic resolution in the z-direction. This method effectively increases the usable thickness of selectively stained samples that can be imaged at a given accelerating voltage by dramatically improving resolution versus unfiltered TEM and increasing signal-to-noise versus zero-loss imaging, thereby expanding the utility of the IVEM to deliver information from within specimens up to 3 microm thick.
Journal of Structural Biology 01/2005; 148(3):297-306. · 3.41 Impact Factor
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ABSTRACT: Data acquisition of cryo-electron tomography (CET) samples described in previous chapters involves relatively imprecise mechanical motions: the tilt series has shifts, rotations, and several other distortions between projections. Alignment is the procedure of correcting for these effects in each image and requires the estimation of a projection model that describes how points from the sample in three-dimensions are projected to generate two-dimensional images. This estimation is enabled by finding corresponding common features between images. This chapter reviews several software packages that perform alignment and reconstruction tasks completely automatically (or with minimal user intervention) in two main scenarios: using gold fiducial markers as high contrast features or using relevant biological structures present in the image (marker-free). In particular, we emphasize the key decision points in the process that users should focus on in order to obtain high-resolution reconstructions.
Methods in Enzymology.
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ABSTRACT: Thesis (Ph. D.)--Catholic University of America, 1945. Bibliography: p. 163-183. Master microform held by: UnM. Microfilm.
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