Focus Documentation

2016workshopc-cina

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

2DX Workshop 2016 The fifth edition of our hands-on workshop on “2D Electron Crystallography” The hand-on workshop on the processing of images from cryo-electron microscopy of 2D crystals of membrane proteins (electron crystallography) will provide the theoretical background and practical applications in advanced image processing algorithms. The workshop will use the 2dx software system and will feature theoretical lectures, hands-on practicals, and give the opportunity to process your own dat…

allspace

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Allspace Umbrella program to calculate internal phase residual in all 17 space groups from data on a single film. Can do the same thing within one film as ORIGTILT does between different films.

allspace1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

ALLSPACE By Vinzenz Unger and Anchi Cheng For a given structure factor list of an untilted crystal, this program is capable of determining its 2-sided plane group from the 17 groups that are possible for chiral molecules. The phases of the symmetry-related reflections have defined relationships. For example, the Friedel mates (h, k, 0) and (-h,-k, 0) should have identical phases (in fact, 0 or 180 °) if there is a 2-fold axis parallel to the z-axis and passing the phase origin of the untilted …

announcementimage2000

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Announcement image2000 Announcement image2000 and plot2k

autocorrl1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

AUTOCORRL By Vinzenz Unger and Anchi Cheng This program calculates an autocorrelation map from a small piece of the filtered image. The central autocorrelation peak is later used to search the cross-correlation map for the position and height of the cross-correlation peaks. Like the cross-correlation (program: TWOFILE) the autocorrelation is calculated in reciprocal space and includes the following separate operations

avrgamphs

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Avrgamphs Program to obtain weighted average phases from merged list.

avrgamphs1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

AVRGAMPHS By Vinzenz Unger and Anchi Cheng AVRGAMPHS is used to average projection data from several images. Because of a scaling error in older versions, version 1.5 or higher should be used. The in- and outputfile are set by FORT1 and FORT2 respectively. The program expects a list of data written by ORIGTILTC/D if the flag on the first input card is set to “T”. Otherwise a format like that produced by ORIGTILTB is assumed. The image data can contain “non-projection” data points that are deri…

backauto

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Backauto Program to calculate the mean radial density (background) of an electron diffraction pattern - any size rectangular area. The new version has options to carry out automatic center determination, and update of the header into the invisible IEXTRA records used by the subsequent program PICKYCOR.

beginninganimageprocessingproject

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Beginning an Image Processing Project A protein project begins with the creation of a folder hierarchy for your images. Begin by creating a folder named for the protein of interest. This folder will be used to hold all images/configuration files related to this particular protein project.

boximage

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Boximage Simple boxing program to create circular or polygonal box with no change in image size.

boximage1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

BOXIMAGE By Vinzenz Unger and Anchi Cheng This program boxes a polygonal area from an input image. The selected area is written out as a new copy of the image without floating, i.e. the image dimensions remain the same and the position of the selected area within the boxed image is unaltered compared to the original. To avoid spikes in the calculated transforms of boxed images, the background is set to the average density around the perimeter of the selected area.

ccunbende

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

CCUNBENDE By Vinzenz Unger and Anchi Cheng CCUNBENDE corrects lattice distortions by reinterpolating the optical densities in the original image based on the positions of the cross-correlation peaks that were determined by QUADSERCH. The input files are the image that is to be corrected (“$image.img” for first pass, “corp1$image.img” for second pass) and a list of the peak positions (CCORDATA: “prof$image.dat”). Only unit cells with peak heights above a chosen fraction of the highest peak (par…

ccunbendk

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Ccunbendk Program unbends the crystal using output from the cross-correlation peak search program, CCORSERCH or its later variants PROFSERCH and QUADSERCH (0, 1, and 2).

changes.htm

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

changes.htm ######0.9.1-7ef0010###### Change Logs 0.9.0 (Developmental Pre-Release) * First implementation of the Focus package. * Contains features like import tool, parallel processing tool. * Various modes can be set depending on the type of the EMproject.

convdeconv

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Convdeconv Program to read in a list of amplitudes and phases and calculate a new list of amplitudes and phases which represent either the convolution or deconvolution of the original.

ctfapply

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

CTFAPPLY By Vinzenz Unger and Anchi Cheng his program corrects the phase data of an individual image for the “contrast transfer function” and astigmatism where applicable. It should be pointed out that CTFAPPLY assumes a square image, which is different from programs involved in the cross-correlation and unbending procedures. Hence, no CTF correction can be applied if the original image is rectangular.

ctfapplyk

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Ctfapplyk Derived from CTFPLOT giving automatic application of CTF to phases input from MMBOX and output ready for ORIGTILT. Program plots spots from MMBOX in reciprocal space and CTF plot of image.

ctffind

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Ctffind This program will attempt to determine the defocus (not yet astigmatism) of any general image whether or not it contains crystalline areas. The program will also apply a CTF correction either by multiplication or division (accompanied by a Wiener filter). Both amplitude and phase contrast with astigmatism can be applied.

ctffind2

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Ctffind2 Determines defocus and astigmatism for images of arbitrary size (MRC format). Astigmatic angle is measured form x axis (same convetiones as in the MRC 2D image processing programs).

ctftilt

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Ctftilt Program determines defocus, astigmatism tilt axis and tilt angle for images of arbitrary size (MRC format). Astigmatic angle is measured from x axis (same conventions as in the MRC 2D image processing programs).

dataflowhere

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Data Flow These pages describe the flow of data through the different programs. File names, formats and contents is listed. 2dx and the underlying MRC programs process and then evaluate images, and extract values for amplitudes and phases of Fourier spots. These are then handed over from one program to the next one, where each program does something to those values (e.g. correcting for the CTF, or merging several values together). Each program thereby unfortunately needs a different type of dat…

emindex.doc

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Emindex.doc Announcement image2000 and plot2k

emrefs.doc

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emrefs.doc References to published papers

explanationofthe2dxgui

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Explanation of the 2dx GUI The upper left pane in the 2dx interface contains all scripts, which will be useful in one round of a sequentially executed workflow. Each script represents an individual processing routine and groups of such scripts may be executed sequentially via multiple selection.

feiextendedmrcformat_not_used_by_2dx

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

FEI extended MRC format (not used by 2dx) Communicated by Remco Schoenmakers, Developer Inspect3D at FEI, on Sept. 5, 2011 The extended MRC format for tomography (as used by FEI; original definition of extended header by Dave Agard and Bram Koster)

fftrans

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Fftrans FFT

fftrans1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

FFTRANS By Vinzenz Unger and Anchi Cheng This program calculates Fourier transforms in either direction (i.e. image --> transform, transform -->image). For a forward transform the real space (image) origin (1,1) is shifted to (1, NY/2+1) and vice versa. The Fourier transform of an image with dimensions NX,NY is (NX/2+1), NY complex values. Each transform pixel therefore contains two parts of information representing the amplitude and phase of the component. Only the in- and output files need t…

frequentlyaskedquestionscontact

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Contact Bug reports Have you encountered any bugs in the software? Please help us identify them using our GITHUB Bug Tracker. Contribute We would be more than happy if you have suggestions and wish to contribute in the development of the software. All you have to do is to sign up on GitHub and contact the developers or submit an issue to the bug-tracker.

histo

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

HISTO By Vinzenz Unger and Anchi Cheng This program produces a listing and a plot of the number of occurences of optical densities within an array of any size. The only input parameter are the starting and end points in x and y, respectively. The type of plot (linear or logarithmic) can also be chosen. To be able to view the listing of the densities a logfile should be created rather than running the program online. An error message “number of out-of-bounds plot errors

home

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Focus 1.1 The current minor release version is 1.1.0. Tutorial Movies A number of movies is available that demonstrate installation, setup, and running FOCUS. The full playlist can be found on YouTube Introductory and Tutorial Movies Hardware recommendations FOCUS requires C++, gfortran, Qt, tcsh and Python. It uses several third-party programs in the background. Several of these don't run under Microsoft Windows. For these reasons, FOCUS only works on LINUX or

image.doc

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

image.doc Some useful documentation for Image Analysis and File Format Programs & Routines

introductiontoelectroncrystallography

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Structure determination from electron micrographs of two-dimensional crystals By Vinzenz Unger and Anchi Cheng The following section provides a general description how to analyze (low-dose) electron micrographs of two-dimensional crystals. In summary the whole procedure can be divided into 5 steps:

label

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Label Program to perform simple manipulations on image files

label1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

LABEL By Vinzenz Unger and Anchi Cheng Label is a general purpose program to perform several useful manipulations. A full list of options can be found in the header of the source code. The following are the options that are used throughout the protocols:

latline

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Latline This is a program to do fitting of curves to either Intensity or F,Phi data for individual layer lines.

latline1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

LATLINE By Vinzenz Unger and Anchi Cheng This program performs fitting of curves to the experimental amplitude and phase data for individual lattice lines. Fitting of curves to a set of scattered input data allows the generation of discrete and uniformly sampled structure factors which can be used to calculate a 3-D density map by Fourier summation. For a more detailed description of the underlining concepts, the readers are encouraged to read the original paper on the program (Agard, 1983) a…

latlineprescal

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Latlineprescal Program to correct image amplitudes for effects of CTF and to calculate appropriate weights for output to LATLINE.

listoffrequentlyaskedquestions

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Frequently Asked Questions My MRC images from Digital Micrograph/eTomo/Other Microscopy Package don't seem to load in 2dx. This can be caused by the “MRC” file not fully being to the standard of the MRC file format loading library provided with the image2000 MRC release. As a result, images in MRC format from eTomo, Digital Micrograph and possibly others can occasionally not be correctly read by the MRC programs that run underneath 2dx.

maketran

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Maketran This program reads in a MTZ file of diffraction amplitudes and phases from the experimentally determined and merged 3-D data from a 2-dimensional crystal, and writes out the transform of the projection of the structure in the direction, at the magnification, and modified by the requested CTF for later use as a reference in cross-correlation with the digitized experimental image.

masktran

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Masktran Program for masking a Fourier transform.

masktrana

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

MASKTRANA By Vinzenz Unger and Anchi Cheng This program is used to mask Fourier transforms. A modified copy of the transform is written out in which only selected parts have non-zero values. The shape and size of the maskholes can be varied as can the mode of operation using either all spots to a given resolution or only those spots for which the (h k) indices are explicitly stated. The most commonly used maskhole shape is a “sharp edged circle” (shape = 1). In this mode the intensities of the…

mergingin2dx_mergein2d

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Merging in 2dx_merge in 2D This processing merges several images into one dataset. This here describes the merging of several non-tilted images into a two-dimensional dataset. Origtiltd: Avramphs: Centric2.exe: 2dx_hklsym.exe: sort: 2dx_hklclean.exe:

mergingin2dx_mergein3d

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Merging in 2dx_merge in 3D This processing merges several images into one dataset. This here describes the merging of several tilted images into a three-dimensional dataset.

mlalgorithmappliedto2dcrystals

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

ML algorithm applied to 2d crystals Maximum likelihood (single micrograph) The parameters used in the maximum likelihood processing are shown in the above GUI. Once the parameters are set, clicking the “maximum likelihood” in the upper left panel starts the process. The obtained reference in each iteration of ML can be viewed by clicking the image name

mmboxa

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Mmboxa Derived from Judy Smith's NNBOX program but with more statistics suitable for more high resolution crystallographic analysis.

mmboxa1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

MMBOXA By Vinzenz Unger and Anchi Cheng This program is used to retrieve and analyze raw phase and amplitude data from a Fourier transform of an image. Setting IOUT≥1 will produce a listing of H, K, amplitude, phase, IQ, background and a dummy column that is later used by CTFAPPLY for the addition of CTF values for each reflection. The logfiles contain valuable statistics, which can help to assess the quality and resolution of the image (see below). If your images require tight boxing of relati…

mmlatref

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Mmlatref Refinement of lattice parameters for maximum amplitude.

mrcdocumentation

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

MRC Dokumentation These documents are taken from the original documentation of the MRC package, as in December 2005. Emrefs.doc References to published papers Plot2k.doc Plot 2000 specifications Image.doc Some useful documentation for Image Analysis and File Format Programs & Routines

mrcfileformatspecifications

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

MRC file format specifications This information is copied from The MRC file header is of 1024 bytes long, organized as 56 LONG words followed by space for 10 80 byte text labels. The full specification, listed in Table 1, is abstracted from the image2000 Announcement of the MRC Image Processing package by Crowther et al., which is currently freely available to academic workers. Use of the programs is described in published papers which …

mrcheaders

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

MRC Headers Headers of the MRC programs, but in the 2DX modified version. Please use these as *rough* guidelines, but check in the actual programs for the current state of your local copy. Padbox Adds data around the edge. Ttboxk Prints out amplitudes & phases in N * N boxes from a Fourier transform, fully corrected for contrast transfer function in tilted image.

mrcprograms

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

MRC programs Guidelines to the original MRC programs LABEL By Vinzenz Unger and Anchi Cheng HISTO By Vinzenz Unger and Anchi Cheng TAPEREDGE By Vinzenz Unger and Anchi Cheng BOXIMAGE By Vinzenz Unger and Anchi Cheng AUTOCORRL By Vinzenz Unger and Anchi Cheng TWOFILE By Vinzenz Unger and Anchi Cheng

mrcsoftware

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

MRC Software Documentation for the MRC program suite Introduction to Electron Crystallography By Vinzenz Unger and Anchi Cheng MRC Headers Headers of the MRC programs, but in the 2DX modified version. Please use these as *rough* guidelines, but check in the actual programs for the current state of your local copy.

origtiltd

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

ORIGTILD By Vinzenz Unger and Anchi Cheng This program serves a number of different purposes such as data merging, calculation of common phase origins or the refinement of tilt parameter. The combination of data from different images requires the determination of their common phase origin ($nprog=1,3 - see following sections). This calculation as well as options for the refinement of several image parameter like the tilt geometry ($nprog=3) and amount of beamtilt ($nprog=1,3) are the most imp…

origtiltk

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Origtiltk Three-dimensional origin, beam tilt and crystal tilt refinement.

padbox

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Padbox Adds data around the edge

parametersdomask

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Domask Switch, deciding if the non-crystal areas in the image should automatically be masked. Domask is a boolean switch that decides if the script Unbend II should automatically mask the non-crystal areas in the image. If this switch is set to “

parametershere

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Parameters Descriptions of the Parameters used in 2dx Initialisation Lattice Magnification and Digitizer Step Size Masking Domask Switch, deciding if the non-crystal areas in the image should automatically be masked. Merging Resolution Zstar Minmax The minimum and maximum grey values for the images. MI Periodogram Phacon Phaori Phase Origin of the dataset for generation of a final map

parametersinitialisation

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Initialization initialization_executable This flag assures that the Initialization script is only executed once. Assuming, 2dx is installed in /usr/local/2dx, your home directory is “~”, and your image processing directory is ~/MyProtein-00/IMAG0012345600

parameterslattice

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Lattice Determine Lattice & Tilt [det_lattice] Switch, deciding if this script should run or not. Lattice Determination Algorithm to run [do_lattice_algorithm] Gives you the option to choose between Findlattice and Getlattice. If realcell unit values have not been obtained, choose

parametersmagnificationanddigitizerstepsize

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Magnification and Digitizer Step Size Magnification (between sample and recording medium) [magnification] The magnification variable defines the nominal magnification of the image. This should be the magnification number that the TEM displayed while recording the image. It is recommended to factor any additional post-magnification factors of the CCD camera or scanner details into the

parametersmasking

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Masking Masking Fourier Radius (To blur the border of the mask) [fourmask] Choose the radius in Fourier Space that will be used for blurring the mask. eg 0.1. Crop into smaller, square image? [mask_crop] Choose if the original size should be kept (n), or if the output image should have smaller dimensions (y).

parametersmerging

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Merging Merging modus Modus of Merging [merge_modus] This allows to define if 2dx_merge should operate in the “2D” or the “3D” modus. In the 2D modus, available data from non-tilted images are merged into a two-dimensional dataset. In the

parametersmi

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Ml Maximum likelihood is an algorithm that finds a set of underlying parameters which maximize the likelihood of the observed particle images. wise The optimization targets include the statistical parameters of translation, rotation, and noise. The algorithm here accomplishes a single particle processing strategy (Fig.), where the unit cells are extracted from 2d crystals. The profile of unbending provides the initial approximate positions of the unit cells.

parametersminma

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Minmax The minimum and maximum grey values for the images. Minimum grey value, and maximum grey value The minimum and maximum grey values specify the upper and lower cut-offs for the images. A micrograph that was digitized on a scanner sometimes has dust pixels that are completely out of range with respect to the majority of the pixels in the image. The same holds for CCD camera images that have hot pixels or dead pixels or suffers from

parametersperiododgram

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Periodogram Generate periodogram? [generatePeriodogram] Periodogram is an algorithm that computes a sharper CTF for better image processing. Setting the periodogram to “yes” creates a sharper CTF through tiling the original image and calculating the power spectrum (PS) for each tile. This allows for higher resolution and better further unbending results.

parametersphacon

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Phacon Phase contrast portion for CTF correction [phacon] allows to specify the amount of Phase Contrast in the image. This information is used for the correction of the Contrast Transfer Function in ctfapply. The Phase Contrast portion is corrected as elastically scattered, and its CTF is calculated as the sinus of the phaseshift of the beam in the sample. The

parametersphaori

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Phaori Phase Origin of the dataset for generation of a final map Phase Origin [phaori] allows to specify the phase origin that should be applied while creating a final map. This phase origin will also be used when merging this dataset to other data from other images. Two registers are provided to specify the phase origin that should be applied for 1) p1 non-symmetrized reconstructions, and 2) for symmetrized reconstruction. These might be different for a tilted sample, if a slightly different…

parametersrealcell

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Real Cell Real-Space Unit Cell and Real-Space Cell Angle realcell and realang define the real-space dimensions of the vectors that describe the unit cell of the protein crystal. This concerns the biological sample, as it swims in the Eppendorf cup. It has (almost) nothing to do with your image. The real-space unit cell dimensions are given in Angstroms, the included angle between the vectors a and b is given in degrees. Bacteriorodopsin for example has real-cell vector lengths of 57.9 Angstrom…

parametersrefine

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Refine Merging Refinement Refine Defocus? [refdefocus] Switch, deciding if defocus should be refined. Refine tilt geometry (only in 3D mode)? [reftiltgeo] Define if during the refinement step also the tilt geometry should be refined. This is done by the MRC program origtilt, which compares the current dataset from one tilted image with the 3D dataset, to find a better tilt geometry. Use with caution, and only on well advanced dataset.

parametersresolution

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Resolution Common Image Processing Upper Resolution Limit [RESMAX] is the resolution limit that should be applied during the image processing, in Angstroem. If for example you are processing a negative stain image, than usually you would want to limit the included resolution to

parametersrotrev

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Rotrev Merging These are all switches allowing to manipulate the Amps and Phases. Rotate +90deg about z? [rot90] Switch, deciding if data should be rotated by 90 deg about z. That means H=-K, K=H. Rotate +180deg about z? [rot180] Switch, deciding if data should be rotated by 180 deg about z. That means H=-H, K=-K.

parametersspidermap

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Spidermap For these parameters to take affect you will need to have the SPIDER software installed. Spider scripts Generate correctly scaled map with SPIDER? (interpolated and sheared multi-unit-cell) [produceSpiderMap] When this is set to “yes”, then a spider script is launched that produces a version of a final map that has a specified scale (mapscale), where the real space dimensions and the angles are correctly interpreted. The final map can also show an adjustable number of unit cells.…

parametersspotlist

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Spotlist The spot list is used to generate the reference for the unbending. SpotList Determination Protect currently existing spotlist [ProtSpotLis] This option allows to decide whether the spotlist generated should be protected from overwriting.

parametersspots

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Spots Source of spots [spot_source] Define where the spotlist should come from. FFT of image is running MMBOX over the FFT of the raw image. The second option creates a full set of all spots within the resolution ranges. The third option does nothing, which only makes sense if the spotlist has been determined before. If not, the third option defaults back to the first one.

parameterssymmetry

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Symmetry Symmetry [SYM] This pull-down menu item allows to specify the symmetry of the 2D crystal sample. For an image of a non-tilted sample, the symmetry can be determined with David Agard's program allspace, using the specific script Get Spacegroup & PhaseOrigin *. This symmetry will be applied during the generation of the symmetrized map, in the specific script *Generate SymMap. An image of a tilted sample has always a p1 symmetry, or no symmetry at all. The symmetry selector can neverthel…

parameterstempkeep

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Tempkeep Keep Large Temporary Files ? [tempkeep] is a switch that decides if temporary files should be kept or deleted.tempkeep allows to opt for keeping large temporary files, thereby filling up more hard-disk space. This may be useful for debugging the image processing, or to study what exactly happens in each image processing step. Once the unbending does satisfying work, the tempkeep switch should be set back to

parameterstilt

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Tilt Tilt Geometry Tilt Geometry [det_tilt] This is a switch, deciding if the tilt angle and tilt axis for this image should be determined, or if the image should instead be considered as non-tilted. It takes some processing time, which is not needed if you already know that the image is from a non-tilted sample. If the switch is set to

parameterstiltgeometry

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Tiltgeometry Variables, defining the tilt geometry The definition of tilt geometry can be found here. ---------- The tilt geometry is defined by 5 variables: TLTAXIS: X-axis -> Tilt axis (on image) [TLTAXIS] Defines the angle from the X-axis to the tilt axis. This is measured on the micrograph. The value is given in grad, and is measured counter-clock wise from the X-axis upwards. TLTAXIS has nothing to do with the 2D crystal.

parameterstreatspotscan

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Treatspotscan Treat SpotScan image? [treatspotscan] a boolean switch allowing to activate a special SpotScan unbending. SpotScan image recording was introduced by Ken Downing ~1992. Instead of exposing the micrograph with a flood beam illumination that covers the entire sample and records the entire micrograph in one go, the beam is condensed into a smaller spot that

parametersttfcorfirst

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

TTFcorfirst is a boolean switch that decides if the TTF should be corrected before or after the unbending TTF correction before unbending (not recommended) ? [ttfcorfirst] allows to choose between two algorithms that correct the Tilted Transfer Function (TTF) either before (yes) or after (no) unbending. The usual procedure would be to first unbend the raw image, and then calculate the FFT of the unbent image. TTBOX can then evaluate the spot amplitudes and phases and their local background un…

parametersttrefine

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

TTrefine Algorithm Selection Only used in the Refine Tilt from SpotSplitting script. Mode of operation [mode] Mode lets you choose things that could be refined. Number of cycles [ncyc] The number of refinement cycles in ttrefine. The values range from 1-1000.

parameterszstar

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Zstar 3D: zstarwin (Window in z* for consideration in phase origin search) [zstarwin] Defines the small vertical window in Fourier space within which the reference data set will be searched for reference data for alignment of a new image. zstarwin should be 2/ALAT, to achieve highest 3D resolution. There is one case, when a larger zstarwin is needed: If you have a reconstruction from non-tilted data only, and then want to add for the first time data from a tilted image, to initiate a 3D recons…

pickauto

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Pickauto Program to integrate spots on a lattice.

pickprofk

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Pickprofk Program for electron diffraction spots integration, using least-squares profile-fitting.

plot2k.doc

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Plot2k.doc Plot 2000 specifications

prepmkmtz

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

PREPMKMTZ By Vinzenz Unger and Anchi Cheng This program is used to convert the fitted data to a format suitable for processing by CCP4 and to automatically reject noisy spots from the data set. The latter is achieved by multiplying the phase error calculated by LATLINE with a constant factor ($reduac) and rejecting those data whose error is ≥90˚ after the multiplication. For instance a factor of 1.5 would result in a rejection of all fitted data that have a phase error of ≥60˚. In its original…

processinganindividualimage

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Processing an individual image With parameters set appropriately you are ready to begin image processing. The scripts on the left represent individual processing steps which need to be performed to fully process an image. Calculate FFT begins image processing by calculating the initial Fourier transform of a tapered version of your original image. Additionally it alters any parameters, which are obviously incorrect in the database. In particular the image side length will be automatically…

processingin2dx_imageofoneimage

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Processing in 2dx_image of ONE image This processing deals only with one single image and is therefore two-dimensional processing. UNBEND II: mmboxa.exe: CORRECT CTF: ctfapply.exe: GENERATE MAP: origtiltd Avramphs: Centric2.exe: F2mtz.exe:

quadserchb

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

QUADSERCHB By Vinzenz Unger and Anchi Cheng This program is a key component in the procedure. It determines the exact positions of the cross-correlation peaks in the real space cross-correlation map, which is obtained by inverse transformation of the cross-correlation transform (“correl$image.fft”) calculated by TWOFILE. The peak positions are written to a file (“prof$image.dat”) which later provides the data for CCUNBEND to calculate the distortion vectors.

quadserck

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Quadserchk Cross-correlation searching program.

refiningalattice

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Refining a lattice The lattice refinement tool can be accessed via the navigator menubar or via the keyboard shortcut: Shift-R. To begin refining select a spot in the Fourier transform on one of the lattice nodes. At the bottom of the window the current point will be displayed as well as its Miller index, generated by the current lattice Information. Initial Miller index guesses might be incorrect, but after definition of the first two or three correct lattice points, they should be accurate en…

refiningthectf

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Refining the CTF The automated defocus search used in 2dx is based entirely on ctffind2 (Grigorieff) and will find the CTF in a majority of cases. However when vibration or other distorting factors destroy parts of the CTF, or in cases where the fit is imprecise, it may be desirable to fit this manually.

rfiltim

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Rfiltim Rotational filtering of images in reciprocal space.

scalimamp3d

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Scalimamp3d Program to scale up image amplitudes to restore the resolution-dependent fall off due to image blurring, radiation damage, temperature factor, static disorder, charging or any other source of power loss.

spotselection

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Spot Selection Spot refinement is needed to generate an initial reference map for cross correlation in the unbending steps. Automatic spot detection is based on the local signal to noise ratio of the peak to the surrounding background. The step arguably requires the highest level of user-interaction due to the subjective nature of the points being considered.

statuspane

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Status Pane The Status Pane in the bottom right corner of the 2dx_image GUI gives information about the current status of the processing of the concerned image. An example is shown here:

stepfive

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Step 5 By Vinzenz Unger and Anchi Cheng Determination of tilt geometries The determination of accurate tilt geometries is important for the buildup of a three-dimensional data set. In praxis, the crystal tilts are usually different from the nominal tilt of the sample holder and deviations of ±5-8˚ are commonly observed. This may just be acceptable for a thin specimen. However, it is strongly recommended to avoid using nominal tilts as starting estimates, especially for thick specimens (>120-1…

stepfour

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Step 4 By Vinzenz Unger and Anchi Cheng Combination of projection data from different images Projection data describe an untilted view of the whole structure projected onto a single section and are comprised of a set of unique reflections with indices (h,k,0). Despite the very limited amount of reciprocal space that needs to be sampled in projection, the data from several images need to be combined because not all structural details are preserved in every electron micrograph. Exactly how many…

stepone

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Step 1 By Vinzenz Unger and Anchi Cheng Optical diffraction, image acquisition and preparation, determination of lattice parameters, calculation of unit cell dimensions and magnification, calculation of a filtered image Optical diffraction Objective:

stepsix

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Step 6 By Vinzenz Unger and Anchi Cheng Merging of 3D data, lattice line fitting and data refinement Building up a 3D data set de novo is very straightforward for specimens that are <100Å thick. In these cases the variations in the molecular transform are relatively slow and can easily be identified even if the initial estimates for defocus, astigmatism and the tilt geometry are quite inaccurate. However, the task gets more demanding as the specimen thickness increases. Because the molecular …

stepthree

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Step 3 By Vinzenz Unger and Anchi Cheng Determination of plane group symmetry CTF-corrected data can always be combined in “plane group p1” (Holser ref), i.e. assuming no symmetry at all. However, if the specimen does exhibit a certain symmetry data handling and refinement becomes a lot easier because symmetry imposes constraints on the molecular transform. For instance, a simple twofold axis of symmetry means that the densities within the unit cell are related by a 180˚ rotation about this a…

steptwo

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:27+00:00

Step 2 By Vinzenz Unger and Anchi Cheng Correction of lattice distortions, extraction of raw image data and correction of phases for the effect of the contrast transfer function (CTF) The steps that are necessary to generate the basic image data are outlined in Fig.4.

symmetrydefinitions

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Symmetry definitions Symmetry definitions used in MRC and 2dx

syncfitp3

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Syncfitp3 Fits SYNC functions to lattice lines produced by MERGHIGH with or without preparation of input data set for plotting version. Includes estimates of errors in SYNC function coeffs and errors in fitted curve.

taperedge

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Taperedge Simple program to taper edges of a rectangular image so that there are no sharp discontinuities which make the Fourier transform contain spikes.

taperedge1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

TAPEREDGE By Vinzenz Unger and Anchi Cheng This program creates smooth density gradients across the edges of an image to avoid spikes in the calculated Fourier transform. The gradients are calculated using the densities on opposite edges of the inputfile (“$image.img”) and a tapered copy of the image (“$image.tap”) is written out. ITAPER/JTAPER designate the total depth along the x and y axes (in pixel) over which the tapering takes place. The maximal allowed setting have been given in the pro…

thefull-screenimagenavigator

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

The Full-Screen Image Navigator The image navigator is designed primarily to provide information about the current image. However, in addition to displaying the image, the current reciprocal lattice and other parameters, the navigator provides the option to manually refine parameters as well. In particular the defocus of the image (and thereby the CTF), the primary and secondary reciprocal lattices and the spot list used to generate initial unbending references can be edited or refined.

tiltgeometrydefinition

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Tilt Geometry Definition An email from Richard Henderson, sent on June 12, 2000 to Henning Stahlberg. TO: Henning Stahlberg FROM: Richard Henderson SUBJ: TTBOX 12-June-2000 Dear Henning, Concerning TTBOX, in order to avoid problems later with remembering how any film has been scanned, I always scan films in the same way - with the number at the top and the edge of the film parallel to the edge of the scanner. TTBOX is written in such a way that the tilt-axis can be described in whatever o…

ttbox

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

TTBOX By Vinzenz Unger and Anchi Cheng The changes in the amount of underfocus across images of highly tilted crystals induce a splitting of the high-resolution spots in the transform due to the effects of the so-called “tilt-transfer function” (TTF) (Shiske 1982, Hayward&Stroud, Henderson et al 1986,1990). An easy way to grasp the basic problem is to recall that a change in defocus across the image will manifest itself as a “breathing” of the Thon ring pattern. Calculating CTF-curves for incr…

ttboxk

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Ttboxk Prints out amplitudes & phases in N * N boxes from a Fourier transform, fully corrected for contrast transfer function in tilted image.

ttboxref

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Ttboxref Refines lattice parameters in image transform of tilted specimen.

ttmask

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:30+00:00

Ttmask Program for masking a Fourier transform, with simultaneous correction for tilted contrast transfer function.

ttmask1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

TTMASK By Vinzenz Unger and Anchi Cheng Changes of the CTF across the image of a highly tilted crystal require a modification to the simple protocol for cross-correlation and lattice straightening to account for the splitting of spots at higher resolutions. In practice this is achieved by replacing MASKTRANA by TTMASK, which is a hybrid between MASKTRANA and TTBOX. TTMASK will correct the transform for the TTF (see TTBOX) before applying a mask. Almost all of the input parameters are identical…

ttrefine

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Ttrefine Refines defocus, astigmatism, tiltaxis and tiltangle in images of thin tilted crystals.

twofile

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Twofile Program to perform simple additions or multiplications on 2 images of same dimensions and type (i.e. both must be real or both complex).

twofile1

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

TWOFILE By Vinzenz Unger and Anchi Cheng This is a general utility program to add or multiply/divide two (complex) images. As for the autocorrelation the cross-correlation between two images can be most easily calculated in Fourier space because the Fourier transform of the cross-correlation is simply the product of the transform (A) of one image and the complex conjugate transform (B*) of the other image. However, the order of the two transforms is not arbitrary in this multiplication. To obt…

twolat

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Twolatt Plots two lattices to see if there is an overlap.

workshop2006

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

Electron Crystallography Workshop 2006 Electron Crystallography Workshop at UC Davis, August 7-11, 2006. Organization: Rena Hill (RMHill@ucdavis.edu) Scientific Organization: Ben Hankamer (B.Hankamer@imb.uq.edu.au) Henning Stahlberg (HStahlberg@ucdavis.edu)

workshop2008

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Workshop 20018 Videos Aims The workshop on membrane protein 2D crystallization and electron crystallography is aimed at PhD students, PostDocs, and beyond, who are interested in the structure determination of membrane proteins (or other 2D protein crystal arrays) by electron crystallography. Topics

workshop2010c-cina

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:28+00:00

Electron Crystallography Workshop 2010 The workshop on membrane protein 2D crystallization and electron crystallography is aimed at everybody (PhD students, PostDocs, group leaders, etc.), who is interested in the structure determination of membrane proteins (or other 2D protein crystal arrays) by electron crystallography.

workshop2012

Anonymous (anonymous@undisclosed.example.com) — 2020-10-22T16:25:29+00:00

NCCR TransCure Hands-On Workshop on Image Processing with 2dx Videos Workshop Program The hand-on workshop on the processing of images from cryo-electron microscopy of 2D crystals of membrane proteins (electron crystallography) will provide the theoretical background and practical applications in advanced image processing algorithms. The workshop will use the 2dx software system, which is based on the MRC programs. Th…