Software tools for molecular microscopy

There are a large number of software tools or software applications that have been specifically developed for the field sometimes referred to as molecular microscopy or cryo-electron microscopy or cryoEM. Several special issues of the Journal of Structural Biology (see references below) have been specifically devoted to descriptions of these applications and several web sites provide partial lists of the software packages and where to obtain them. This article is being created to provide a starting point for a more complete list and up-to-date distribution information of all of the software of interest to the cryoEM community. The software tools described here have been loosely and somewhat arbitrarily organized into several categories as follows:

General packages: Packages that offer a comprehensive set of tools to permit the analysis of data in several classes of structural problems.

Specific packages: Packages that offer a comprehensive set of tools to permit the analysis of data in a single class of structural problem. For example packages specifically focused on objects with helical, icosahedral, crystalline symmetry, etc. Application tools: Packages that offer a tool or a set of tools to permit the analysis of data in one or more class of structural problems. These have generally been developed to manage one specific step in the structural analysis, for example CTF correction, particle picking etc.

Visualization and modeling tools: Packages that facilitate the interpretation, analysis or presentation of the results.

Utilities: General utilities like file format conversion etc.

General packages:
Packages that offer a comprehensive set of tools to permit the analysis of data in several classes of structural problems.

Bsoft
 * Bsoft is a collection of programs and a platform for development of software for image and molecular processing in structural biology. Problems in structural biology are approached with a highly modular design, allowing fast development of new algorithms without the burden of issues such as file I/O. It provides an easily accessible interface, a resource that can be and has been used in other packages. Several workflows such as for single particle analysis and tomography are supported with parameter exchange as well as the ability to do distributed processing across heterogeneous clusters of computers. Several tools are available for modeling structures as atomic coordinates or larger scale models for the interpretation of large molecular complexes.


 * Version: 1.5.0 Contact: Bernard_Heymann at nih.gov  OS support: Unix (Mac OS X, IRIX, Linux, AIX, Solaris, Tru64), VMS Image format support: BioRad, Brix, CCP4, Digital Instruments, Digital Micrograph, EM, Goodford, GRD, Imagic, JPEG, MFF, Image Magick, MRC, PIC, PIF, PNG, Spider, Suprim, TIFF, RAW Cost: Free


 * Publications:
 * Heymann, J. B. (2001) Bsoft: image and molecular processing in electron microscopy. Journal of Structural Biology 133 (2-3): 156 - 69.
 * Heymann, J. B., and Belnap, D. M. (2007) Bsoft: Image processing and molecular modeling for electron microscopy. Journal of Structural Biology 157: 3 - 18.
 * Heymann, J. B., Cardone, G., Winkler, D. C. and Steven, A. C. (2007) Computational resources for cryo-electron tomography in Bsoft. Journal of Structural Biology in press.

Cyclops
 * Cyclops is a new computer program designed as a graphical front-end that allows easy control and interaction with tasks and programs for 3D reconstruction of biological complexes using cryo-electron microscopy. It was designed for straightforward implementation in grid architectures. As a front-end to a collection of programs it provides a common interface to other programs, thus enhancing the usability of the suite and the productivity of the user.


 * Version: Unknown Contact: plaisier @chem.leidenuniv.nl  OS support: Unknown  Image format support: Unknown  Cost: Free/GPL license
 * Publications:
 * Plaisier, J. R., Jiang, L., and Abrahams, J. P. Cyclops: New modular software suite for cryo-EM. Journal of Structural Biology In Press, Corrected Proof.

EMAN (and now alsoEMAN2)
 * A suite of scientific image processing tools aimed primarily at single particle reconstruction. This is a technique for determining the 3-D structure of a molecule or macromolecular assembly from thousands to hundreds of thousands of noisy images of individual molecules, generally collected on a transmission electron microscope. EMAN's focus is on providing state of the art single particle reconstruction methods automated to the greatest extent possible. The goal is to permit even novice users to be able to reconstruct macromolecular structures with high veracity and at high resolution. It also has a variety of tools for more generic image processing, useful for electron tomography, 2-D crystallography and helical reconstructions.


 * EMAN consists of a ~100,000 line C++ library with bindings to the popular Python programming language. It offers literally hundreds of different scientific image processing algorithms including Fourier processing, real-space filters, 3-D reconstruction, projection, etc. In EMAN2, all user-level programs, including GUI programs, are written in Python, permitting the advanced user to easily customize aspects of the package. EMAN is funded by the NIGMS through grant R01GM080139.


 * Version: 1.8
 * Contact: sludtke@bcm.edu
 * OS support: Linux, OSX, most Unix variants, Windows (GUI and utilities only)
 * Image format support: HDF5, SPIDER, MRC/CCP4, IMAGIC, PIF, ICOS, VTK, PGM, Amira, Xplor, Gatan DM2/DM3, TIFF, Scans-a-Lot, LST, PNG, V4L, JPEG
 * Euler angle conventions: EMAN, SPIDER, IMAGIC, MRC, SGI, CHIMERA
 * Cost: Free/Open Source
 * Publications to Cite:
 * Ludtke, S. J., Baldwin, P. R., and Chiu, W. (1999). EMAN: Semiautomated Software for High-Resolution Single-Particle Reconstructions. Journal of Structural Biology 128, 82-97.
 * Tang, G., Peng, L., Baldwin, P. R., Mann, D. S., Jiang, W., Rees, I., and Ludtke, S. J. EMAN2: An extensible image processing suite for electron microscopy. Journal of Structural Biology. 157:38-46.

EMStudio
 * A comprehensive data processing system for single-particle electron microscopy. It provides functions for automatic & manual particle selection, image CTF correction, particle alignment, particle classification, model reconstruction, and map/PDB structural analysis.  Its graphic user interface creates an informative and friendly computing environment to facilitate and expedite the data processing.


 * Version: Beta Contact: jzchen @brandeis.edu OS support: Linux, Mac OS X, MS Windows  Image format support: MRC, TIFF  Cost: Free to academic community

IMAGIC
 * A high end environment for the analysis of images, spectra and other multi-dimensional data-sets. IMAGIC-5's software package is aimed at processing (massive) data sets from electron, light and raster-tunnelling microscopes, computer tomographs, FT-IR spectrometers and other signal collecting devices.


 * Version: -5 Contact: imagic@ImageScience.de  OS support: Most platforms  Image format support: IMAGIC-5 Cost:  Starting at: non profit: 6,500USD/5.100 Euro
 * Publications:
 * Van Heel M., Harauz G., Orlova E.V., Schmidt R. and Schatz M. (1996) A new generation of the IMAGIC image processing system, J. Struct. Biol. 116: 17-24.

iplt
 * The Image Processing Library & Toolbox is an open-source project primarily aimed the electron microscopy community, with particular emphasis on 2D electron crystallography. It consists of several modular class libraries written in C++, each exposing a majority of their interface to Python, upon which the processing logic is build. Easy extendability has been a prime focus for the framework design, and we welcome any contribution from the community.


 * Version: Unknown Contact: ansgar dot philippsen at unibas dot ch  OS support: Linux, Unix, Irix, Mac OS X Image format support: MRC, CCP4, TIFF, PNG, DM3, SPIDER Cost: Free/GPL
 * Publications:
 * Philippsen, A., Schenk, A. D., Signorell, G. A., Mariani, V., Berneche, S., and Engel, A. Collaborative EM image processing with the IPLT image processing library and toolbox. J Struct Biol. 2007 Jan;157(1):28-37 link
 * Philippsen A, Schenk AD, Stahlberg H, Engel A. IPLT: image processing library and toolkit for the electron microscopy community. J Struct Biol. 2003 Oct-Nov;144(1-2):4-12. link

MDPP
 * The Micrograph Data Processing Program (MDPP) is a fully-featured general purpose image processing package originally written to support research in structural biology requiring electron microscopy and image processing.


 * Version: v06.100 Contact: smithp01 @med.nyu.edu OS support: Mac OS/Unix/VMS Image format support: Many Cost: Free
 * Publications:
 * Smith, P. R., and Gottesman, S. M. (1996). The Micrograph Data Processing Program. Journal of Structural Biology 116, 35-40.

MRC IMAGE PROCESSING PACKAGE

The MRC image processing package has been developed over many years, with contributions by many authors. It comprises software for image processing of 2D crystals, analysis of electron diffraction patterns, helical diffraction and single particle analysis, especially particles with icosahedral symmetry. It also includes visualization software for displaying and manipulating images in many ways.


 * Contact: jms@mrc-lmb.cam.ac.uk OS support: DEC/Tru64, LINUX, UNIX, IRIX, Mac OS X
 * Image Format support: MRC Cost: free to academic users:
 * Publications:
 * Crowther, R.A., Henderson, R. and Smith, J.M. (1996). MRC Image Processing Programs. Journal of Structural Biology 116, 9-16.
 * Smith, J.M. (1999) XIMDISP - A visualization tool to aid structure determination from electron microscope images. Journal of Structural Biology 125, 223-228.

SPARX
 * SPARX (Single Particle Analysis for Resolution eXtension) is a new image processing environment with a particular emphasis on transmission electron microscopy (TEM) structure determination. It includes a user interface that provides a complete graphical programming environment with a novel data/process-flow infrastructure, an extensive library of replace.py scripts that perform specific TEM-related computational tasks, and a core library of fundamental C++ image processing functions. In addition, SPARX relies on the EMAN2 library and CCTBX, the open source computational crystallography library from PHENIX.


 * Version: 0.3 Contact: sludtke@bcm.tmc.edu, pawel.a.penczek@uth.tmc.edu  OS support: Most platforms  Image format support: Most formats  Cost: Free/Open Source
 * 'Publications to Cite:''
 * Hohn, M., T., Tang, G., Goodyear, G., Baldwin, P.R., Huang, Z., Penczek, P.A., Yang, Ch., Baldwin, P.R., Adams, P., Ludtke, S.J.: SPARX, a new environment for cryo-EM image processing. J. Struct. Biol., 157:47-55, 2007.
 * Baldwin, P.R., Penczek, P.A.: The transform class in SPARX and EMAN2. J. Struct. Biol., 157:250-261, 2007.

Spider


 * SPIDER (System for Processing Image Data from Electron microscopy and Related fields) is an image processing system for electron microscopy. Contains numerous operations for: 3D reconstruction, averaging of single particle macromolecule specimens, multivariate statistical classification of images, and electron tomography.  Computational modules are written in Fortran and the visualization GUI is written in C. Developed and maintained since 1978.


 * Version: 15.06+ Contact: spider@wadsworth.org  OS support:Unix (Linux, Irix, AIX, Mac OS X)  Image format support: SPIDER, CCP4, Emispic, MRC, PDB, Raw Cost: Free (GNU General Public License)
 * Publications:
 * Baxter, W. T., Leith, A., and Frank, J. (2007) SPIRE: The SPIDER reconstruction engine. Journal of Structural Biology 157, 56-63.
 * Frank, J., Radermacher, M., Penczek, P., Zhu, J., Li, Y., Ladjadj, M., and Leith, A. (1996). SPIDER and WEB: Processing and Visualization of Images in 3D Electron Microscopy and Related Fields. Journal of Structural Biology 116, 190-199.
 * Yang, C., Penczek, P. A., Leith, A., Asturias, F. J., Ng, E. G., Glaeser, R. M., and Frank, J. (2007) The parallelization of SPIDER on distributed-memory computers using MPI. Journal of Structural Biology 157, 240-249.

Suprim
 * A flexible, modular software package intended for the processing of electron microscopy images. The system consists of a set of image processing tools or filters,written in the C programming language, and a command line style user interface based on the UNIX shell. The pipe and filter structure of UNIX and the availability of command files in the form of shell scripts eases the construction of complex image processing procedures from the simpler tools.


 * Version: 5.5 Contact: amisoftware@scripps.edu  OS support: Unix Image format support: MRC/suprim Cost: Free
 * Publications:
 * Schroeter, J. P., and Bretaudiere, J.-P. (1996). SUPRIM: Easily Modified Image Processing Software. Journal of Structural Biology 116, 131-137.

Xmipp
 * "X-Window-based Microscopy Image Processing Package", is a specialized suite of image processing programs primarily aimed at obtaining the three-dimensional reconstruction of biological specimens from large sets of projection images acquired by transmission electron microscopy.


 * Version: 1.1 Contact: xmipp@cnb.uam.es  OS support: UNIX Image format support: SPIDER, TIFF Cost: Free/Open Source
 * Publications:
 * C.O.S. Sorzano, R. Marabini, J. Velazquez-Muriel, J.R. Bilbao-Castro, S.H.W. Scheres, J.M. Carazo, A. Pascual-Montano (2004) XMIPP: a new generation of an open-source image processing package for Electron Microscopy. Journal of Structural Biology 148(2) pp. 194-204.

Specific packages:
Packages that offer a comprehensive set of tools to permit the analysis of data in a single class of structural problem. For example packages specifically focused on objects with helical, icosahedral, crystalline symmetry, etc.

Two-dimensional crystals
2dx
 * A software system designed as a user friendly, platform-independent software package for electron crystallography. 2dx assists in the management of an image-processing project, guides the user through the processing of 2D crystal images, and provides transparence for processing tasks and results. Algorithms are implemented in the form of script templates reminiscent of c-shell scripts.
 * 2dx builds upon the MRC programs, which were extended by additional functions to interface with the GUI of 2dx, and to implement the optionally automatic processing.


 * Version: 2.1.0 Contact: HStahlberg@ucdavis.edu or BRGipson@ucdavis.edu  OS support: Mac OS X (PPC and Intel), Linux (64bit and 32bit) Image format support: MRC, TIFF, CCP4 Cost: Free/GNU General Public License
 * Publications:
 * Bryant Gipson, Xiangyan Zeng, Zi Yan Zhang and Henning Stahlberg (2007). 2dx – User-friendly image processing for 2D crystals. Journal of Structural Biology 157(1), 64-72.
 * Ludovic Renault, Hui-Ting Chou, Po-Lin Chiu, Rena M. Hill, Xiangyan Zeng, Bryant Gipson, Zi Yan Zhang, Anchi Cheng, Vinzenz Unger, and Henning Stahlberg (2006). Milestones in electron crystallography. J. Comp.-Aided Molec. Design 20, 519-527.

Icosahedral viruses
AUTO3DEM
 * An automation system designed to accelerate the computationally intensive process of three-dimensional structure determination from images of vitrified icosahedral virus particles. With minimal user input and intervention, AUTO3DEM manages the flow of data between the major image reconstruction programs, monitors the progress of the computations, and intelligently updates the input parameters as the resolution of the model is improved.


 * Version: 2.02 Contact: sinkovit@sdsc.edu  OS support: Linux, UNIX Image format support: PIFCost: Free/BSD license
 * Publications:
 * Yan X., R.S. Sinkovits, and T.S. Baker (2007) AUTO3DEM - an automated and high throughput program for image reconstruction of icosahedral particles. J. Struct. Bio. 157:73-82.

Helices
[IHRSR]
 * The iterative helical real space reconstruction (IHRSR) algorithm can recontruct helical filaments under conditions when traditional Fourier–Bessel approaches sometimes fail. For example when there is disorder or heterogeneity present, when the specimens diffract weakly, or when Bessel functions overlap.


 * Version: Unknown Contact: Edward H. Egelman, University of Virginia, egelman@virginia.edu   OS support: Linux  Image format support: SPIDER  Cost: Free
 * Publications:
 * Egelman, E. H. The iterative helical real space reconstruction method: Surmounting the problems posed by real polymers. Journal of Structural Biology 2007 Jan;157(1):83-94

Phoelix
 * A set of procedures and algorithms for helical processing that we refer to as the PHOELIX package. The package was developed to provide a time-efficient and semiautomated method for determining a three-dimensional density map from a specimen with helical symmetry. The procedures which are part of  PHOELIXn are drawn from the original MRC helical processing suite with extensions principally developed using the SUPRIM image processing package. The package in its current form has been optimized for the processing of actomyosin filaments but has been modified and applied to other helical structures.


 * Version: 1.3 Contact: amisoftware@scripps.edu  OS support: Unix Image format support: MRC/suprim Cost: Free/Open source
 * Publications:
 * Carragher, B., Whittaker, M., and Milligan, R. A. (1996). Helical Processing Using PHOELIX. Journal of Structural Biology 116, 107-112.
 * Whittaker, M., Carragher, B. O., and Milligan, R. A. (1995). PHOELIX: a package for semi-automated helical reconstruction. Ultramicroscopy 58, 245-259.

Ruby-Helix
 * Set of programs for the analysis of “helical” objects with or without a seam. Ruby-Helix is built on top of the Ruby programming language and is the first implementation of asymmetric helical reconstruction for practical image analysis. It also allows easier and semi-automated analysis, performing iterative unbending and accurate determination of the repeat length..


 * Version: Unknown Contact: mkikkawa@em.biophys.kyoto-u.ac.jp  OS support: Fedora, Mac OS X Image format support: MRC Cost: Free/Open Source
 * Publications:
 * Kikkawa, M. A new theory and algorithm for reconstructing helical structures with a seam.   J Mol Biol. 2004 Oct 29;343(4):943-55
 * Metlagel, Z., Kikkawa, Y. S., and Kikkawa, M. Ruby-Helix: An implementation of helical image processing based on object-oriented scripting language. Journal of Structural Biology 2007 Jan;157(1):95-105

StokesLabProcedures
 * Image Processing Software for Helical Crystals: outlines, procedures and hints for using the MRC helical processing software which has since been modified by several groups including Chikashi Toyoshima, Nigel Unwin, David DeRosier and David Stokes.


 * Version: - Contact: stokes@saturn.med.nyu.edu  OS support: Unix Image format support: MRC Cost: Free/Open Source
 * Publications:

Single particles
Frealign
 * Frealign provides algorithms optimized for the efficient refinement of three-dimensional reconstructions and correction for the contrast transfer function of the microscope in the determination of macromolecular structures by single particle electron microscopy.


 * Version: 7 Contact: niko @brandeis.edu  OS support: Linux, IRIX, OSF, Mac OS X Image format support: MRC, Spider Cost: Free
 * Publications:
 * Grigorieff, N. (2007). FREALIGN: High-resolution refinement of single particle structures. Journal of Structural Biology 157, 117-125.

PFT3DR
 * PFT3DR is package for determining orientations and origins of imaged particles and computing a three-dimensional reconstruction from the images and their assigned orientations and origins. The programs are enhanced versions of the PFT algorithm developed by Baker and Cheng (2000) and the Fourier Bessel reconstruction algorithm of Crowther et al. (1970).  The original adaptations of the PFT3DR programs were specific to icosahedral viruses, but were enhanced to support nearly all symmetries found in biological particles.  Other enhancements are described on the PFT3DR web site.


 * Version: 2  Contact: David_Belnap at byu.edu   OS support: Unix (Linux, Mac OS X, etc.), VMS  Image format support: same as Bsoft (see above)  Cost: Free, Open Source
 * Publications:
 * Baker, T. S. and R. H. Cheng. (1996). A model-based approach for determining orientations of biological macromolecules imaged by cryoelectron microscopy. J. Struct. Biol. 116, 120-130.
 * Crowther, R. A., L. A. Amos, J. T. Finch, D. J. De Rosier, and A. Klug. (1970) Three dimensional reconstructions of spherical viruses by Fourier synthesis from electron micrographs.
 * Fuller, S. D., S. J. Butcher, R. H. Cheng, and T. S. Baker. (1996) Three-dimensional reconstruction of icosahedral particles--the uncommon line. J. Struct. Biol. 116, 48-55.
 * Bubeck, D., D. J. Filman, N. Cheng, A. C. Steven, J. M. Hogle, and D. M. Belnap. (2005) The structure of the poliovirus 135S cell entry intermediate at 10-angstrom resolution reveals the location of an externalized polypeptide that binds to membranes. J. Virol. 79, 7745-7755.

Tomography
IMOD
 * IMOD is a set of image processing, modeling and display programs used for tomographic reconstruction and for 3D reconstruction of EM serial sections and optical sections. The package contains tools for assembling and aligning data within multiple types and sizes of image stacks, viewing 3-D data from any orientation, and modeling and display of the image files. It includes a complete graphical user interface for generating tomograms, combining tomograms from tilt series taken around two axes, and stacking tomograms from serial sections.


 * Version: 3.7 Contact: mast at colorado dot edu  OS support: Linux, Windows, Mac OS X Image format support: MRC, TIFF Cost: Free
 * Publications:
 * Kremer, J. R., Mastronarde, D. N., and McIntosh, J. R. (1996). Computer Visualization of Three-Dimensional Image Data Using IMOD. Journal of Structural Biology 116, 71-76.
 * Mastronarde, D. N. (1997) Dual-axis tomography: an approach with alignment methods that preserve resolution. Journal of Structural Biology 120, 343-352.

protomo
 * Protomo is a suite of programs and shell scripts primarily developed for electron tomography. It offers routines for preprocessing micrographs, CTF-correction of images of untilted and tilted specimens, marker-free alignment of tilt series, and 3D reconstruction, besides some general image processing functionality.


 * Version: 1.1 Contact: protomo@electrontomography.org  OS support: Linux Image format support: Most formats Cost: Free
 * Publications:
 * Winkler, H. (2007) 3D reconstruction and processing of volumetric data in cryo-electron tomography. Journal of Structural Biology 157, 126-137.
 * Winkler, H. and Taylor, K. A. (2006) Accurate marker-free alignment with simultaneous geometry determination and reconstruction of tilt series in electron tomography. Ultramicroscopy 106, 240-254.
 * Winkler, H. and Taylor, K. A. (2003) Focus gradient correction applied to tilt series image data used in electron tomography. Journal of Structural Biology 143, 24-32.
 * Taylor, K. A., Tang, J., Cheng, Y., and Winkler, H. (1997) The use of electron tomography for structural analysis of disordered protein arrays. Journal of Structural Biology 120, 372-386.

SerialEM
 * SerialEM is a program to acquire tilt series for electron tomography on Tecnai and newer JEOL microscopes. It uses an approach based on prediction of specimen position during the tilt series from the position at previous tilts. With this method, it achieves both the robustness of the older approach to tilt series acquisition (track and focus at every tilt) and the speed of the newer precalibration approach. It provides a complete interface for camera control and image acquisition, viewing, and saving. It includes a low dose mode for tracking and focusing away from the area of interest, energy filter control, acquisition of tilt series using a montage of overlapping frames, and a navigator module for mapping the grid and returning to selected locations.  It supports Gatan and Tietz CCD cameras.


 * Version: 2.6  Contact: mast at colorado dot edu  OS support: Microsoft Windows  Image format support: MRC  Cost: Free for academic use.
 * Publications:
 * Mastronarde, D. N. (2005). Automated electron microscope tomography using robust prediction of specimen movements. Journal of Structural Biology 152, 36-51.

TOM
 * The Tomography Toolbox is a collection of functions that extend the capability of the MATLAB® (+ Image Procesing Toolbox) numeric computing environment. The toolbox supports a wide range of functions for tomography.


 * Version: not versioned Contact: tom@biochem.mpg.de  OS support: Linux, Microsoft Windows Image format support: EM,MRC,Spider Cost: Free
 * Publications:
 * Nickell, S., Forster, F., Linaroudis, A., Net, W. D., Beck, F., Hegerl, R., Baumeister, W., and Plitzko, J. M. (2005). TOM software toolbox: acquisition and analysis for electron tomography. Journal of Structural Biology 149, 227-234.

UCSF Tomography
 * UCSF Tomography is an integrated software suite that provides full automation from tomographic data collection to real-time reconstruction as well as automated acquisition of random conical data sets. This software was implemented based upon a novel approach in which the compustage tilting is modeled as geometric rotation. The spatial movement of the sample as a result of stage tilting can be predicted based upon previously collected tomographic images. Therefore, there is no need to collect tracking and focusing images during the entire tomographic data collection. A significant dose saving can thus be achieved and is critical in collecting cryo tilt series. Real-time reconstruction is achieved by calculating a weighted back-projection on a small Linux cluster (five dual-processor computer nodes) concurrently with the UCSF tomography data collection running on the microscope’s computer, and using the fiducial-marker free alignment data generated during the data collection process. The real-time reconstructed 3D volume provides users with immediate feedback to fully asses all aspects of the experiment ranging from sample choice, ice thickness, experimental parameters to the quality of specimen preparation. An on-site scheme was developed for random conical data collection where tracking and focusing are performed at the same location as the final conical tilt images. Lower magnifications combined with short exposure are used to substantially reduce dose and to allow larger tilt steps. The system also includes a feature for montaging untilted images to ensure that all of the particles in the tilted image may be used in the reconstruction.


 * Version: v7.7.3J  Contact: agard@msg.ucsf.edu  OS support: Windows 2000, XP Image format support: MRC Cost: Free for academic use
 * Publications:
 * Zheng, Q. S., Braunfeld, M. B., Sedat, J. W., and Agard, D. A. (2004). An improved strategy for automated electron microscopic tomography. Journal of Structural Biology 147, 91-101.
 * Zheng, S. Q., Keszthelyi, B., Branlund, E., Lyle, J. M., Braunfeld, M. B., Sedat, J. W., and Agard, D. A. UCSF tomography: An integrated software suite for real-time electron microscopic tomographic data collection, alignment, and reconstruction. Journal of Structural Biology 157, 138-147.
 * Zheng, S. Q., Kollman, J. M., Branlund, M. B., Sedat, J. W., and Agard, D. A. Automated acquisition of electron microscopic random conical tilt sets. Journal of Structural Biology 157, 148-155

EM3D
 * EM3D is a software application designed to analyze and visualize electron microscope (EM) tomography data. It is especially for cellular and molecular biologists. From a tilt series of 2D electron micrographs taken at many tilt angles with respect to the electron beam, this program can then perform auto-align and quickly render those data into a lucid 3D model, which allows you to perform object rotating for viewing. In addition, EM3D also provides analysis tools for quantify structural information from the models, including their moments, proximity relationships, and spatial reliability.  And all of these functions can be executed with a very intuited graphic user interface. EM3D is available free-of-charge for most computer operation systems, including Mac OS X for PowerPC or Intel, UNIX, Linux, and Windows. EM3D is being developed in the laboratory of Dr. U. J. McMahan, Professor of Neurobiology and of Structural Biology at Stanford University School of Medicine.


 * Version: v2.0  Contact: contact@em3d.stanford.edu  OS support: Windows XP, Linux, Mac OS X for PowerPC or Intel Image format support: Unknown Cost: Free for academic use.


 * Publications:
 * EM3D's scheme for the automatic alignment of tilt-images is described. Ress, D., Harlow, M.L., Schwarz, M., Marshall, R.M., and U.J. Mc Mahan. Automatic acquisition of fiducial markers and alignment of images in tilt series for electron tomography. J. Electron Microscopy 48: 277-287, 1999
 * EM3D is first used for exposing celluar architecture at macromolecular resolution. Harlow, M.L., Ress, D., Stoschek, A., Marshall, R.M. and McMahan, U.J. The architecture of active zone material at the frog's neuromuscular junction. Nature 409:479-484, 2001
 * EM3D's method for creating optimal isodensity surface models is described. Ress, D., Harlow, M.L., Marshall, R.A., and McMahan, U.J. Optimized Method for Isodensity Surface Models Obtained with Electron Microscope Tomography Data. Engineering in Medicine and Biology Society, 2003. Proceedings of the 25th Annual International Conference of IEEE. pp. 774-777, 2003.
 * EM3D is used to examine the architecture of the postsynaptic density at brain synapses. Petersen, J.D., Chen, X., Vinada, L., Dosemeci, A., Lisman, J.E., and Reese, T.S. Distribution of postsynaptic density (PSD)-95 and Ca2+/calmodulin-dependent protein kinase II at the PSD. J. Neuroscience, 23: 11270-11278, 2003.
 * EM3D approach to model generation is detailed. Ress, D.B., Harlow, M.L., Marshall, R.M. and McMahan, U.J. Methods for generating high-resolution structural models from electron microscope tomography data. Structure: 12 (10):1763-1774, 2004.

Application tools:
Packages that offer a tool or a set of tools to permit the analysis of data in one or more class of structural problems. These have generally been developed to manage one specific step in the structural analysis, for example CTF correction, particle picking etc.

Data Acquisition
Leginon
 * Leginon is a system designed for automated collection of images from a transmission electron microscope. Instruments supported: FEI Tecnai series TEM, Tietz and Gatan CCD cameras. On August 1, 2007, Leginon 1.4 was released as open source under the Apache License, Version 2.0 and is Free to download.


 * Version: 1.2 Contact: amisoftware@scripps.edu  OS support: Linux/PCImage format support: MRC Cost: Free for academic use.
 * Publications:
 * Suloway, C., Pulokas, J., Fellmann, D., Cheng, A., Guerra, F., Quispe, J., Stagg, S., Potter, C. S., and Carragher, B. (2005). Automated molecular microscopy: The new Leginon system. Journal of Structural Biology 151, 41-60.

SerialEM
 * SerialEM is a program to acquire tilt series for electron tomography on Tecnai and newer JEOL microscopes. It uses an approach based on prediction of specimen position during the tilt series from the position at previous tilts. With this method, it achieves both the robustness of the older approach to tilt series acquisition (track and focus at every tilt) and the speed of the newer precalibration approach. It provides a complete interface for camera control and image acquisition, viewing, and saving. It includes a low dose mode for tracking and focusing away from the area of interest, energy filter control, acquisition of tilt series using a montage of overlapping frames, and a navigator module for mapping the grid and returning to selected locations.  It supports Gatan and Tietz CCD cameras.


 * Version: 2.6  Contact: mast at colorado dot edu  OS support: Microsoft Windows  Image format support: MRC  Cost: Free for academic use.
 * Publications:
 * Mastronarde, D. N. (2005). Automated electron microscope tomography using robust prediction of specimen movements. Journal of Structural Biology 152, 36-51.

UCSF Tomography
 * UCSF Tomography is an integrated software suite that provides full automation from tomographic data collection to real-time reconstruction as well as automated acquisition of random conical data sets. This software was implemented based upon a novel approach in which the compustage tilting is modeled as geometric rotation. The spatial movement of the sample as a result of stage tilting can be predicted based upon previously collected tomographic images. Therefore, there is no need to collect tracking and focusing images during the entire tomographic data collection. A significant dose saving can thus be achieved and is critical in collecting cryo tilt series. Real-time reconstruction is achieved by calculating a weighted back-projection on a small Linux cluster (five dual-processor computer nodes) concurrently with the UCSF tomography data collection running on the microscope’s computer, and using the fiducial-marker free alignment data generated during the data collection process. The real-time reconstructed 3D volume provides users with immediate feedback to fully asses all aspects of the experiment ranging from sample choice, ice thickness, experimental parameters to the quality of specimen preparation. An on-site scheme was developed for random conical data collection where tracking and focusing are performed at the same location as the final conical tilt images. Lower magnifications combined with short exposure are used to substantially reduce dose and to allow larger tilt steps. The system also includes a feature for montaging untilted images to ensure that all of the particles in the tilted image may be used in the reconstruction.


 * Version: v7.7.3J  Contact: agard@msg.ucsf.edu  OS support: Windows 2000, XP Image format support: MRC Cost: Free for academic use
 * Publications:
 * Zheng, Q. S., Braunfeld, M. B., Sedat, J. W., and Agard, D. A. (2004). An improved strategy for automated electron microscopic tomography. Journal of Structural Biology 147, 91-101.
 * Zheng, S. Q., Keszthelyi, B., Branlund, E., Lyle, J. M., Braunfeld, M. B., Sedat, J. W., and Agard, D. A. UCSF tomography: An integrated software suite for real-time electron microscopic tomographic data collection, alignment, and reconstruction. Journal of Structural Biology 157, 138-147.
 * Zheng, S. Q., Kollman, J. M., Branlund, M. B., Sedat, J. W., and Agard, D. A. Automated acquisition of electron microscopic random conical tilt sets. Journal of Structural Biology 157, 148-155

TOM
 * The Tomography Toolbox is a collection of functions that extend the capability of the MATLAB® (+ Image Procesing Tooolbox) numeric computing environment. The toolbox supports a wide range of functions for tomography.


 * Version: not versioned Contact: tom@biochem.mpg.de  OS support: Linux, Microsoft Windows Image format support: EM,MRC,Spider Cost: Free
 * Publications:
 * Nickell, S., Forster, F., Linaroudis, A., Net, W. D., Beck, F., Hegerl, R., Baumeister, W., and Plitzko, J. M. (2005). TOM software toolbox: acquisition and analysis for electron tomography. Journal of Structural Biology 149, 227-234.

Particle Selection
Signature
 * A particle selection system for molecular electron microscopy. It applies a hierarchical screening procedure to identify molecular particles in EM micrographs. The user interface of the program provides versatile functions to facilitate image data visualization, particle annotation and particle quality inspection. The system design emphasizes both functionality and usability.


 * Version: Unknown Contact: jzchen @brandeis.edu  OS support: Linux, Mac OS X, MS Windows Image format support: MRC, TIFF  Cost: Free to academic community
 * Publications:
 * Chen, J. Z., and Grigorieff, N. (2007). SIGNATURE: A single-particle selection system for molecular electron microscopy. Journal of Structural Biology 157, 168-173.

SwarmPS Developer's page
 * A specialised graphical user interface designed to streamline the process of particle selection from electron micrograph data sets. It provides implementations of both template matching and edge detection algorithms, has an intuitive and easy to use interface, and can be used to obtain useful results on most data sets that are generated in electron microscopy. Currently users can expect to select about 1000-4000 particles per hour of interaction.


 * Version: 0.9.2 Contact: d.woolford @imb.uq.edu.au  OS support: Linux Image format support: Most image formats Cost: Free for academic use under a software license agreement.
 * Publications:
 * Woolford, D., Ericksson, G., Rothnagel, R., Muller, D., Landsberg, M. J., Pantelic, R. S., McDowall, A., Pailthorpe, B., Young, P. R., Hankamer, B., and Banks, J. (2007). SwarmPS: Rapid, semi-automated single particle selection software. Journal of Structural Biology 157, 174-188.

CTF Estimation and Correction
ACE
 * A completely automated algorithm for estimating the parameters of the contrast transfer function (CTF) of a transmission electron microscope. A MATLAB implementation of the algorithm, called ACE, is freely available. See also author's website: http://graphics.ucsd.edu/~spmallick/research/ace/index.html)


 * Version: 2.3.1 Contact: amisoftware@scripps.edu OS support: Unknown Image format support: MRC Cost: Free/Open Source
 * Publications:
 * Satya P. Mallick, Bridget Carragher, Clinton S. Potter, David J. Kriegman "ACE: Automated CTF Estimation" Ultramicroscopy, Volume 104, Issue 1, August 2005, Pages 8-29.

ctffind ctftilt
 * Programs for finding CTFs of electron micrographs


 * Version: 3.1/1.2 Contact: niko @brandeis.edu OS support: Linux, IRIX, OSF Image format support: MRC Cost: Free
 * Publications:
 * Mindell, J. A., and Grigorieff, N. (2003). Accurate determination of local defocus and specimen tilt in electron microscopy. Journal of Structural Biology 142, 334-347.

TOMOCTF
 * Package for CTF determination and correction in electron cryotomography


 * Version: 1.0 Contact: jjfdez at ual.es ; jjfdez at mrc-lmb.cam.ac.uk OS support: Linux, OSF Image format support: MRC Cost: Free for academic users
 * Publications:
 * Fernandez, J. J., Li, S. and Crowther, R. A. (2006). CTF determination and correction in electron cryotomography. Ultramicroscopy 106, 587-596.

Resolution Estimation
FSC
 * Program to calculate the Fourier Shell Correlation (FSC) of two 3D volumes.


 * Version: Unknown Contact: info@ImageScience.de   OS support: Linux, WindowsImage format support: IMAGIC, Spider, CCP4, MRC, etc. Cost: Free
 * Publications:
 * George Harauz and Marin van Heel (1986). "Exact filters for general geometry three dimensional reconstruction". Optik, 78, 146–156.
 * Marin van Heel and Michael Schatz (2005) "Fourier shell correlation threshold criteria". Journal of Structural Biology, 151, 250-262.

RMEASURE
 * A computational method that allows the measurement of the signal-to-noise ratio and resolution of a three-dimensional structure obtained by single particle electron microscopy and reconstruction. The method does not rely on the availability of the original image data or the calculation of several structures from different parts of the data that are needed for the commonly used Fourier Shell Correlation criterion. Instead, the correlation between neighboring Fourier pixels is calculated and used to distinguish signal from noise.


 * Version: - Contact: niko @brandeis.edu  OS support: Linux, IRIX, OSF, Mac OS X Image format support: MRC, Spider  Cost: Free
 * Publications:
 * Duncan Sousa and Nikolaus Grigorieff (2007). Ab initio resolution measurement for single particle structures. Journal of Structural Biology 157, 201-210.

Denoising
TOMOAND
 * Package for noise reduction in electron cryotomography with Anisotropic Nonlinear Diffusion.


 * Version: 1.0 Contact: jjfdez at ual.es ; jjfdez at mrc-lmb.cam.ac.uk OS support: Linux, OSF Image format support: MRC Cost: Free for academic users
 * Publications:
 * Fernandez, J. J. and Li, S. (2003). An improved algorithm for anisotropic nonlinear diffusion for denoising cryotomograms. Journal of Structural Biology 144, 152-161.
 * Fernandez, J. J. and Li, S. (2005). Anisotropic nonlinear filtering of cellular structures in cryoelectron tomography. Computing in Science and Engineering 7(5) 54-61.
 * Fernandez, J. J., Li, S. and Lucic, V. (2007). Three-dimensional anisotropic noise reduction with automated parameter tuning. Application to electron cryotomography. Lecture Notes in Computer Science (in press).

Visualization and modeling tools:
Chimera
 * Molecular modeling package with many density map capabilities. Supports fitting atomic models in maps, interactive segmentation, coarse modeling, measuring and coloring of density maps for elucidating structures of large molecular assemblies. The map capabilities were designed for analyzing electron microscope single particle reconstructions and tomography. Includes many methods for analysis of atomic resolution molecular models and multiple sequence alignments.


 * Version: 1.2415  Contact: chimera-users@cgl.ucsf.edu  OS support: Windows, Macintosh, Linux, IRIX and Tru64 Image format support: MRC, CCP4, SPIDER, BRIX, SITUS, PIF, others Cost: Free for academic use
 * Publications:
 * Goddard TD, Huang CC, Ferrin TE. (2007). Visualizing density maps with UCSF Chimera. Journal of Structural Biology 157(1):281-7.

EMfit
 * Program for fitting atomic models into electron microscopy maps. Fitting criteria includes the sum of densities at atomic sites, the lack of atoms in negative or low density, the absence of atomic clashes between symmetry-related positions of the atomic structure, and the distances between identifiable features in the map and their positions on the fitted atomic structure, etc.


 * Version: 5.0 Contact: mr@indiana.bio.purdue.edu  OS support: Linux, AIX. MAP format support: TSB ASCII EM, XPLOR ASCII. Cost: Free / Open Source
 * Publications:
 * Rossmann, M. G., R. Bernal, S. V. Pletnev (2001). Combining electron microscopic with X ray crystallographic structures. Journal of Structural Biology 136, 190-200.
 * Rossmann, M. G.(2000). Fitting atomic models into electron microscopy maps. Acta Crystallogr. D56, 1341-1349.

RIVEM
 * Program developed to project electron density of virus radially onto a sphere that is then presented as a stereographic diagram. Features that constitute the viral surface can be simultaneously represented in terms of atoms, amino acid residues, potential charge distribution, and surface topology.


 * Version: 3.2 Contact: xc@purdue.edu  OS support: Linux, AIX. MAP format support: XPLOR ASCII. Cost: Free
 * Publications:
 * Xiao, C., Rossmann, M. G. (2007). Interpretation of electron density with stereographic roadmap projections. Journal of Structural Biology 158, 182-187

Situs
 * Package for the modeling of atomic resolution structures into low-resolution density maps e.g. from electron microscopy, tomography, or small angle X-ray scattering..


 * Version: 2.3 Contact: situs@biomachina.org  OS support: Windows/Cygwin, Macintosh, Linux, IRIX. Image format support: MRC, CCP4, SPIDER, XPLOR, ASCII. Cost: Free / Open Source (GNU GPL)
 * Publications:
 * Wriggers, W., Milligan, R. A., McCammon, J. A. (1999). Situs: A Package for Docking Crystal Structures into Low-Resolution Maps from Electron Microscopy. Journal of Structural Biology 133, 185-195.
 * Wriggers, W., Chacón P., Kovacs J., Tama F., and Birmanns, S. (2004). Topology Representing Neural Networks Reconcile Biomolecular Shape, Structure, and Dynamics. Neurocomputing 56, 365-379.

UROX
 * An interactive tool for fitting atomic models into electron microscopy reconstructions (or into envelopes derived from small-angle X-ray or neutron scattering). The correlation between the electron density of the atomic models and the map is calculated and updated as a model is moved on the graphical display with the help of the mouse. Calculations are performed in reciprocal-space and the symmetry of the reconstruction is taken into account. The computations are fast and an entire EM reconstruction can be used.


 * Version: 1.1.1 Contact: xsiebert@gmail.com  OS support: Linux MAP format support: EZD, CCP4, MRC Cost: Free
 * Publications:
 * Navaza, J., Lepault, J., Rey, F. A., Alvarez-Rua, C., Borge, J. (2002). On the fitting of model electron densities into EM reconstructions: a reciprocal-space formulation. Acta Cryst. D38, 10.

Utilities:
em2em
 * Program to convert images from/to formats typically used in the EM community.


 * Version: Unknown Contact: imagic@ImageScience.de  OS support: Most platforms  Image format support: Most formats  Cost: Free
 * Publications:

References:

 * Advances in Computational Image Processing for Microscopy, JSB volume 116 1996
 * Chiu W, Baker ML, Jiang W, Dougherty M, Schmid MF. Electron cryomicroscopy of biological machines at subnanometer resolution. Structure. 2005 Mar;13(3):363-72.
 * Frank, Joachim (2006). Three-Dimensional Electron Microscopy of Macromolecular Assemblies: Visualization of Biological Molecules in Their Native State, 2nd edition, Oxford University Press. ISBN 0-19-518218-9.
 * Software tools for macromolecular microscopy, Journal of Structural Biology, Volume 157, Issue 1, Pages 1-296 (January 2007). Edited by Bridget Carragher, Clinton Potter and Fred Sigworth