midas(1) General Commands Manual midas(1) NAME midas - manual image alignment for MRC files. SYNOPSIS midas [options] image_file [transform_file] DESCRIPTION Midas allows one to assess and adjust the alignment between pairs of images. It is typically used to align images from serial sections and to correct errors in a pre-alignment of tilt series images by cross- correlation. It can also be used to adjust the alignment between adja- cent pieces in a montaged image. There are interactive controls for adjusting translation, rotation, magnification (scale), and stretch along an axis. Linear transformations are used to apply the transla- tion, rotation or distortion. This means that the same amount of translation, rotation or distortion happens everywhere in an image. A nonlinear transformation, or warping, can be added by marking selected points and locally aligning images at those locations. All transformation parameters can be adjusted by dragging a point on the image with the mouse. The help menus describe the functions of each mouse button, as well as the operation of the various controls and hot keys. The controls also have tooltips with brief summaries of their function. Midas also allows one to adjust the brightness and contrast of each image independently to ease comparisons. The program can produce a copy of the image stack with these individual contrast changes applied (but not with the transformations applied). The input image files for Midas must be MRC files. They can be byte, integer, or real images, and can also be RGB images, which will be dis- played in grayscale. The latter option can be used to align model con- tours displayed on an image by snapshotting the Zap window in 3dmod and creating a color MRC stack with tif2mrc. Information on using Midas with a high-DPI monitor is provided in sec- tion 1.13 of the IMOD Guide. Alignment modes Midas maintains a list of transformations, one for each section in the image file. At any given time, one section is the "current" section, it is displayed with its transformation applied to it, and any adjust- ments in alignment modify that transform. Another section is the "ref- erence" section to which the current section is being aligned. Other aspects of the way that transforms are treated depend on which mode Midas is operating in: local, global, or reference. In local alignment mode, each image section is usually aligned to the previous section. The transformations in this mode are sometimes referred to as "xf" transforms, which are suitable only for aligning one image to another. The program xftoxg is used to convert xf transforms into the "xg" transforms that are suitable for transforming the whole image stack into alignment. In this mode, Midas displays the reference section without transforming it, and any changes in transfor- mation are applied only to the transform of the current section. In global alignment mode, each image section is effectively aligned to the entire image stack. The transformations are of the global (xg) type, ready to use to transform all of the images into alignment. There are two differences in program operation that achieve this: first, the reference section's transformation is applied whenever it is displayed; second, when you modify the transform of the current sec- tion, the transforms of all following sections are changed by the same amount. Global alignment may be suitable for aligning small sets of sections because of its simplicity. However, it is not recommended for larger numbers of sections (more than 20-30). Local alignment followed by the use of xftoxg is preferable because it will automatically keep the aligned images from drifting out of the field of view. It will also preserve trends in the data; for example, if you align using features that gradually get smaller, global alignment may eliminate this size change, while local alignment will preserve it, even if every transform over the series includes a size change. If you do want to use global alignment mode, you may want to start in the middle of the image stack and work forward from there. Then, turn off the "Keep Curr-Ref Diff" box, make the middle section be the refer- ence again and make the previous section be the current section. Turn the "Keep Curr-Ref Diff" box back on again and work backwards to the beginning of the stack. Reference alignment mode is used to align each image in a stack to a single image that may be in another image file. The other image file is specified with the -r option. In this mode, advancing or backing up by a section changes only the current section unless you turn on the "Keep Curr-Ref Diff" option; with that option off, the reference sec- tion is changed only by typing a new number into its text box. If your reference section happens to be in the same file as the images being aligned to it, then you can either specify that file as both the refer- ence and the image file, or start Midas in local alignment mode then turn off the "Keep Curr-Ref Diff" box and set the reference and current sections independently. In global or reference alignment mode, a menu option can be used to transform a model file using the transformations available for each section, just as with imodtrans. Warping Images Warping can introduced by adding a set of points, referred to as con- trol points, and applying an appropriate shift at each point. This warping is in addition to whatever linear transformation has been set. To start adding points, turn on "Add/edit warp points". You can then add a point with the middle mouse button and shift images into align- ment at that location with either the left mouse button, the X and Y translation buttons, or the arrow keys. The shift at the point is shown on the lines with the X and Y translation buttons, while the translation component of the linear transformation is shown on a sepa- rate line. Until there are at least four points, shifting a control point will just change the linear transformation. With one point, it will shift the whole image; with two points, shifting at one point will rotate and magnify the image to keep it stationary at the other point; with three points, shifting at one point will stretch the image to keep it aligned at the other two points. You can thus use three points in this way to set a linear transformation. If the linear transformation is already set, you should probably add four or more points before shifting any of them. Once there are at least four points, you can no longer turn off the option to edit warp points because the linear translation is fixed. Make the control points widely spaced and distributed over as much of the area as possible. The program will warp the image inside the region bounded by the control points but has to use extrapolated trans- formations outside that region. A control point is considered to be fixed on the reference image. You can thus add the point at a recognizable feature on the reference image, then shift the current image to center that feature on the point. You can change the fixed location of the point by clicking with the right mouse button. The currently active point is shown in yellow and other points are shown in red. Click with the left mouse button to select the nearest point as the active point. There is a menu entry and a hot key for deleting the current point. Warping transformations can be specified only in local or reference alignment mode. Xftoxg can convert them to global transformations. They can be used interchangeably with linear transforms in Xfprod- uct(1), Xfinverse, Newstack, Blendmont, Xfmodel, and serial tomogram joining. Storage of Transforms The transformation for the current section is automatically stored on the internal list of transforms when you change to a different section. Before changing sections, you can restore the transform to the stored value using an entry in the Edit menu. There are also menu entries to reset to the unit transform (no translations, rotations, etc.) and to store the current transform on the internal list. You can use the lat- ter to save a transform that you like before trying to improve on it. If the name of a file of transforms is entered after the image filename when first starting Midas, then Midas will read transforms from this file, and images will initially be displayed with these transforms. This file then becomes the default file into which Midas will save transforms. Transforms can be saved into a different file with the File menu entry "Save As". After doing so, that file becomes the default file for successive save operations. In either case, the first time that Midas saves to a file, it will make a backup of an existing file by that name by adding ~ at the end. If the name of a transform file that does not yet exist is entered on the command line, Midas will create and save into that file. Using Global Rotations and Cosine Stretching Midas can rotate all images by a common amount and allow you to adjust transformations in the rotated images. This ability, invoked with the -a option, can be used to make the tilt axis vertical when aligning a tilt series. When the program is started in this mode, there is a con- trol for adjusting the global rotation angle. There is also a check box to constrain translations with the mouse to the X direction, which is usually the only direction that needs fixing in a tilt series pre- aligned by correlation. This rotation provides a convenient way to look at tilted images but should not affect the alignment; the trans- formations output by the program are independent of the global rotation angle. However, the X and Y translations that are displayed and adjusted in Midas are relative to the rotated images and will change with the angle. Global rotations can be used in local or global align- ment mode. Midas can also apply cosine stretching of images if a file of tilt angles is provided with the -t option. It will stretch the current image by the ratio of the cosines of the current and reference image tilt angles. The stretch is always along the X axis after applying a global rotation, so invoking this option turns on the global rotation mode. With such a stretch, it should be easier to find the shift that aligns the images. If stretched images do not line up well, even when looking just at features on one Z plane, then an offset can be applied to the tilt angles. As with global rotation, this stretching does not affect the alignment transforms output by the program, but the X trans- lation that is displayed in the program does depend on the stretching. Cosine stretching cannot be used in global or reference alignment mode. Aligning Chunks of Sections Midas has another mode of operation that can be used to align tomograms of serial sections. This is referred to as "chunk" alignment because sections are considered in chunks, where a chunk corresponds to all of the slices from one tomogram. In this mode, all of the sections in a chunk are given the same transformation alignment, so there is only one transformation per chunk. This mode is a variant of the local align- ment mode, with the additional constraint that the alignment is always between adjacent pairs of chunks. When adjusting the transformation between a pair of chunks, you can view any pair of sections from the two chunks, and you are also free to scroll through the slices in a chunk to discern trends in position. Activate this mode by specifying the number of sections in each chunk with the -c option. To get one transformation per tomogram, do not divide the slices from the top and bottom of tomogram into separate chunks. Fixing Montage Overlaps When the pieces of a montage need to be shifted into registration with each other, Blendmont uses the image displacements in the overlap zones between adjacent pieces to determine how to shift the pieces. Sometimes Blendmont does not determine a displacement correctly, lead- ing to erroneous registration of the pieces. To use Midas to correct the displacements, start Midas with the -p option followed by the name of the piece list file. After the montaged image file name, in place of a transformation file, enter the name of the edge correlation dis- placement (.ecd) file produced by Blendmont. You can then display and adjust the overlap between any pair of adjacent pieces on any section. Overlap zones are referred to as edges and are of two types: X edges between pieces in a horizontal row, and Y edges between pieces in a column. X edges are numbered sequentially from left to right, first in the bottom row, then in the next row, etc. Y edges are numbered from bottom to top in columns, with the leftmost column being the first one. The edge number controls, just below the section controls, allow one to select any given edge by this number, but they also allow you to see and select an edge by the X and Y frame number of the lower piece (the one to the left of an X edge, or below a Y edge). To find edges that need fixing, it is usually more convenient to use the buttons provided for selecting one of the edges with the worst displacement errors. When two pieces are displayed, their X and Y translation values corre- spond to the image displacement at that edge, originally measured by Blendmont and possibly adjusted within Midas. The displacements for all of the edges are used to determine the optimal shifts of the piece. When these shifts are applied to the pieces, they produce displacements at all of the edges, displacements that may or may not match the indi- vidual displacements measured from the images there. A mismatch at an edge is a displacement error, which has X and Y components. Large dis- placement errors arise when the displacement measured at an edge does not align the images there. Manually aligning images at an edge with a large error will thus reduce the error there and also at other edges involving the same pieces. If there is only one edge with a mismeasured displacement, then solving for the shifts using all of the other edges while leaving out the one in question will often imply a perfectly good displacement at that edge. The difference between this displacement and the measured one is called the "leave-out error", and pushing the "Apply Leave-out Error" button will adjust the edge to the displacement implied by all of the other edges. If an edge shows a visible image mismatch that appears to correspond to the leave-out error, then pushing this button is a useful shortcut for aligning the images by hand. One must be careful to examine both of the edges with the top two errors when the erroneous edge involves a corner piece in the montage, a not infrequent occurrence. In this case, the error will be equally divided between the two edges of the corner piece, even though there is an erroneous measurement at only one of them. For similar reasons, a 2 by 2 montage with an erroneous displacement will spread the error equally among all four edges. If the montage consists of only one row or one column, all errors will be zero regardless of whether the displacements have been measured cor- rectly. Visual examination of the edges is the only way to detect and correct errors in the measured displacements. When there are more than 10 pieces, the program can find a more robust solution for the shifts between pieces that can give less weight to or eliminate edges that have obviously erroneous displacement. This is referred to as robust fitting. Its main value in Midas is to make the edges with bad diplacements show up with particularly high errors. The degree of robustness is controlled by a criterion value with a default value of 1. With the default value, some edges that have good dis- placements may be downweighted inappropriately and thus show up as hav- ing a high error even though they are well-aligned. A higher criterion will prevent this from happening but may fail to identify some bad edges as outliers (values up to about 1.5 should be tried). If an edge with a good alignment shows up as having a large error with robust fitting enabled, use the following strategy: 1) Continue to fix or exclude other edges with high errors. The error of the well-aligned edge may drop as other errors around it are resolved. 2) If the well-aligned edge still has a high error after other edges with high errors have been fixed, examine all of the edges around the two pieces below and above this edge. You may find a small misalign- ment. 3) If you still can not find a misaligned edge, raise the fitting cri- terion until the error becomes normal, or ignore the large error. Note that if you are going to use the robust fitting option in Blendmont, you would need to set the criterion to this same value to prevent a misalignment across this edge. OPTIONS Midas accepts some Qt options (such as -style) and the following options. -g Start Midas in global alignment mode. The default is local alignment mode. -r filename Load a reference image from a file with the given filename. This reference image can be used for aligning an image stack to the single reference image. -rz section This option will select the section number to be used when load- ing in a reference image with the -r option. The default value is the first section which is numbered 0. -c list of sizes Determine alignment between chunks of sections. Enter a comma- separated list of the number of sections in each chunk. The list should add up to the total number of sections in the image file. When chunks are entered in this way, the reference or current section can be set to any section within a chunk. -cs list of sizes Determine alignment between chunks of sections from samples near the boundaries between chunks (e.g., tomograms corresponding to physical sections). Enter a comma-separated list of the number of sections in each sample, where the first and last chunk have only one sample and other chunks have two samples. The list should add up to the total number of sections in the image file. When chunks are entered in this way, the reference and current section are constrained to stay within a sample. -B factor Bin images by the given factor when reading them in. The trans- lation component of the transformation shown and adjusted in the program will be smaller by this amount. Translations will be scaled down when read in from a file and scaled back up when written out to file. Errors shown when fixing montage overlaps will also be smaller by the binning factor. Entering this option overrides automatic binning specified with the environ- ment variable MIDAS_BIN_TO_TARGET (use "man imodenv" or "imod- help" - Other Man Pages - IMOD Environment Variables for details). -p filename Load piece coordinates for a montaged image file from the given filename and enter montage fixing mode. The final argument on the command line would ordinarily be the .ecd file produced by Blendmont, but it is possible to open a montage without such a file. -C size Set the cache size to the given number of sections. Sections are read from the file as needed, intensity-scaled to be stored as bytes, and kept in a cache. By default, the cache is limited to about 1 gigabyte, or at least big enough for 4 sections. If your file has more than this number of pixels, and you have more memory available, you can use this option to provide enough cache for all sections. If you have less than this amount of memory available and your file has more pixels than there is memory, you should enter an appropriate size to avoid disk swap- ping. -s min,max Scale the image intensities when reading from the file, so that the range from min to max will be scaled to range from 0 to 255 when the section is stored in the program. -b size Set the initial size of the blocks for transforming the current image. The larger the block size, the faster the display, but quality will suffer, especially for images that are rotated sub- stantially. A size of 0 will turn on image interpolation. -a angle Display all images with a global rotation specified by angle. When the program is started with this option, the angle can be adjusted if necessary with a spin box control. -o filename Save transforms to a file with the given filename instead of to the file that transforms were read from. -e number When fixing montage overlaps, show the given number of buttons with the highest errors (between 2 and 10). -O 2 letters Specify the colors for the previous and current image in overlay mode. The colors must be two of r g b c m y which stand for red, green, blue, cyan (green + blue), magenta (red + blue), and yellow (red + green). The two letters must specify different channels; e.g., rc will work and rm will not. The default over- lay colors are magenta and green (mg). -D Run in debug mode, which means do not run the program in the background. -q Suppress reminder to rebuild edge functions when exiting after fixing montage edges. FILES The linear transformation files that Midas uses are in the following format. The files are in ascii format with one line for each image in the image stack. Each line contains 6 numerical parameters that are used to calculate the linear transformation. A11 A12 A21 A22 DX DY Where for each pixel with the coordinates (X, Y) a new location (X', Y') is given by the equations below (Xc, Yc is the center of the image). X' = A11 * (X - Xc) + A12 * (Y - Yc) + DX + Xc Y' = A21 * (X - Xc) + A22 * (Y - Yc) + DY + Yc Warping transformation files contain a linear transformation of this form plus the set of control points for each section. Their format is described in the libiwarp library documentation. HISTORY Midas (Manual Image Deformation and Alignment System) was first written in Fortran for a MicroVax with Parallax graphics by David Mastronarde. Jim Kremer then wrote a similar program in C for the SGI using first IrisGL, then X-windows, then OpenGL and Motif. This was originally named Midas, but was renamed to Manali after Mastronarde ported the Midas program from the Vax to the SGI. For the next version of Midas, Mastronarde modified Kremer's Manali to incorporate desirable features of the old Midas (such as manipulation with the mouse), to implement new features (such as caching, ability to zoom large images down, abil- ity to read integer or real mode files, and treatment of a transform file the way a model file is treated in 3dmod), and to provide com- pletely new controls, menus, and help windows. In the latest version, the graphical interface was rewritten in Qt. SEE ALSO xftoxg, imodtrans, newstack, blendmont, 3dmod, tif2mrc BUGS Email bug reports to mast at colorado dot edu. IMOD 4.11.0 midas(1)