Boulder Laboratory for 3-Dimensional Electron Microscopy of Cells
MTLENGTHS(1) MTLENGTHS(1)
NAME
mtlengths - compute lengths of MTs in a model
SYNOPSIS
mtlengths
DESCRIPTION
MTLENGTHS computes the lengths, in microns, of contours in
IMOD models, and produces a list giving the object number and contour
length for each contour. This list may be used to generate a
histogram of lengths with a program such as GENHSTPLT. One may
optionally add a certain amount of length to each contour at its
high Z end, so that the length calculation will include the
thickness of every section in which a contour appears.
If the reconstruction is from serial sections that were significantly tilted
during microscopy, the program can adjust for these tilts given the proper
information. Prepare a file in which the first line shows the Z value and
the tilt of the first tilted section (or of the first section, if that one
was tilted), and each successive line shows the Z value and tilt for each
section on which tilt was changed. Z values should occur in ascending
order.
If the contours are mainly perpendicular in the model (e.g.,
microtubules in good cross-section in tilted images), then this simple
adjustment for tilting is adequate. However, if trajectories are
oblique, the program can do a more complex adjustment for the tilting
if it is given information about the location of the tilt axis in the
images. This information consists of the angle of the axis in raw
images (i.e., the angle of rotation from the vertical to the tilt
axis), and optionally the list of transformations applied to align the
raw images. When the program adjusts for obliqueness, there is a
potential ambiguity about the sign of the tilt angles. It thus
computes lengths both with the tilt angles as given and with their signs
inverted. It reports the fraction of line segments that come out
longer with the angles as given than with the angles inverted. This
fraction is referred to as the fraction that are "better", because it
is assumed that the tilting was done in the direction that reduced the
obliqueness of the fibers by as much as possible. This fraction is
reported for each range of Z values at a single tilt angle, and
overall. If the fraction better is less than 0.5, one should rerun the
program, selecting the option to invert tilt angles. On this run, the
angles will be considered to be specified as the negative of the angles
in the tilt file, and the fraction of segments that are longer with the
angles as specified will be greater than 0.5.
These adjustments for tilting are irrelevant for a tomographic
reconstruction.
The program can also be provided with an object containing a contour
that serves as a length marker. It will report the length of this
contour both without and with the correction for obliqueness, and with
the length given for both polarities of the tilt angle.
If one corrects for obliqueness, there is also an option to produce a
set of transformations that can be applied to produce the model that
would have been obtained if the sections had not been tilted.
Imodtrans or Xfmodel can be used to transform the model.
The output of the program depends on whether one is correcting for
obliqeness. If one is not, the output consists of the object number and
the length for each MT. With obliqueness correction, the output consists
of the object and contour number, the length uncorrected for obliqueness,
the length with angles as specified, and the length with angles inverted.
On the final run, with the polarity of angles selected as inverted if
necessary, the desired lengths will be in the fourth column.
Mtlengths uses the PIP package for input (see the manual page for pip)
and can take input interactively if correction for obliqueness is not
required. The following options can be specified either as command line
arguments (with the -) or one per line in a command file or parameter file
(without the -):
-model OR -ModelFile File name
Input model file.
-output OR -OutputFile File name
Output file for lengths. The output file may be omitted for output to
terminal or log file.
-tilt OR -TiltFile File name
File with tilt information in the format described above.
-extra OR -ExtraLength Floating point
Fraction of a section to add at the high Z end of each contour to account
for section thickness. The default is 0.
-magnification OR -Magnification Floating point
Magnification of film images, or 1 if CCD images. This option is required
if there is no scale in the model header.
-scale OR -ScaleDigitized Floating point
Microns per pixel at which film was digitized, or pixel size in microns
for CCD images. This option is required if there is no scale in the model
header.
-section OR -SectionThickness Floating point
Section thickness in nanometers. This option is required if there is no
scale in the model header.
-exclude OR -ExcludeObjects List of integer ranges
Objects to exclude from length measurement.
-marker OR -MarkerObject Integer
Object number of length marker for measuring overall spindle length.
-correct OR -CorrectObliques
Correct lengths of oblique MTs when sections are tilted.
-axis OR -AxisAngle Floating point
Rotation angle from the vertical to the tilt axis in the raw images. This
entry is required when correcting for obliqueness.
-align OR -AlignmentTransforms File name
File with transforms used to align the images. This entry is optional and
is relevant only when correcting for obliqueness.
-invert OR -InvertAngles
Change the sign of the tilt angles when adjusting lengths of oblique
fibers.
-untilt OR -UntiltTransforms File name
Output file for transforms that will correct model for tilted sections.
-param OR -ParameterFile Parameter file
Read parameter entries as keyword-value pairs from a parameter file.
-help OR -usage
Print help output.
-StandardInput
Read parameter entries from standard input.
For compatibility with older command files, the program will also
accept input interactively if started with no command line arguments,
but it will not be able to correct for obliqueness.
The interactive entries to the program are:
Name of model file
Name of file with tilt information, or Return if none
IF the model has no scaling information in its header, make the
following three entries:
Magnification of negatives (without any commas)
Scale, in microns per pixel, at which negatives were digitized
Section thickness in nanometers
Amount to add to the high Z end of each object in order to account
for section thickness. For example, enter 1 to allow a section's
worth of length for each section that an object appears in.
Name of output file in which to place list of lengths, or Return for
output to the terminal
List of IMOD objects to exclude from the output, or Return to output
all objects.
HISTORY
Written by David Mastronarde, 1/27/90; modified for IMOD 4/24/97;
incorporated obliqueness correction and switched to PIP input, 9/4/03