mtffilter(1) General Commands Manual mtffilter(1)NAMEmtffilter - filter by inverse of MTF and general Fourier filterSYNOPSISmtffilter [options] input_file [output_file]DESCRIPTIONMtffilter originated as a program for restoring contrast in camera images by multiplying them by the inverse of the camera's modulation transfer function (MTF), but has evolved to do general-purpose filter- ing in Fourier space as well as to apply some other specialized fil- ters. It can apply a low pass filter to reduce high frequency noise, as well as a high pass filter to eliminate low frequencies. Any combi- nation of these filters may be applied. In fact, the program provides all of the options that other IMOD programs do for specifying a general Fourier filter. Because images are automatically padded to dimensions suitable for taking an FFT, there are no restrictions on image size, unlike with Enhance. This program can filter either real-space images in 2D planes, real-space images in 3D or 3D Fourier transforms in 3D. The filter functions produced by these options can be visual- ized with the program Filterplot; see that man page for a full description of their effects. A specialized filter can be applied to perform dose weight-filtering of cryoEM images, particularly ones from tilt series. The filter is as described in Grant and Grigorieff, 2015 (DOI: 10.7554/eLife.06980) and the implementation follows that in their "unblur" program. At any fre- quency, the filter follows an exponential decay with dose, where the exponential is of the dose divided by 2 times a "critical dose" for that frequency. This critical dose was empirically found to be approx- imated by a * k^b + c, where k is frequency; the values of a, b, c in that paper are used by default but can be modified with the-criticaloption. This filter function is applied for all frequencies (complete attenuation above the "optimal dose" is no longer considered appropri- ate). A variety of options are described below for providing dose information to the program. This filter cannot be combined with the other Fourier filters. However, when dose weighting is activated, the MaximumInverse, InverseRolloffRadiusSigma, and LowPassRadiusSigma options will simply be skipped and have no effect, in order to leave the standard mtffilter command file runnable if dose weighting options are inserted into it with a template. The program can also apply a Wiener-like deconvolution filter that matches what is implemented in Warp (Tegunov and Cramer, 2019, Nat. Methods 16, 1146^a1152) and IsoNet (Y. T. Yiu, et al., 2022, Nat. Comm. 13, 6482). The filter compensates for attenuations by the microscope contrast transfer function (CTF), using an assumed signal-to-noise ratio that starts high and falls off at a specified rate. This filter cannot be combined with the other Fourier filters. The program can also apply a fourth kind of filter that has been found useful for reducing fringe effects in EM images taken with a phase plate. This filter is specified by a cutoff radius, and exponential power, and an amplification factor, the amount by which it amplifies low frequencies relative to high ones. It goes from 1 at zero fre- quency to a floor of 1/ amplificationFactor at high frequencies and falls 1/e of the way to the floor at the cutoff radius. A higher power increases the sharpness of the falloff. This filter is referred to here as a low-frequency amplifier filter. It is selected by entering the -amplifier option plus either the cutoff radius (with -cutoff) or parameters of the phase plate imaging (with -phase). It cannot be used together with the low-pass and high-pass filter options. The program can also apply a filter in one dimension, in the X-direc- tion only, and specifically can apply an R-weighted 1-D filter such as is used in back-projection. This R-weighted filter cannot be used together with inverse MTF filtering. Simply multiplying by the inverse of an MTF would amplify noise too much, so the inverse MTF filter is shaped by three parameters. The first and most important is the maximum inverse value, which limits how high the inverse can become. The other two parameters are a cutoff frequency at which to start a Gaussian rolloff of the inverse back to 1.0, and the sigma value for this Gaussian rolloff. The default values for these parameters (listed below) are based on limited experimenta- tion and are fairly conservative. All of these parameters together will keep the inverse filter from amplifying high frequency noise. The low pass filter's role is to filter out those high frequencies. If both filters are used, there are potentially 4 different frequency ranges: 1) From 0 to the frequency at which the inverse reaches its maximum, the filter is actually the inverse of the MTF, 2) From there to the cutoff frequency for the inverse rolloff, the filter equals the maximum inverse, 3) Beyond this cutoff frequency, the filter progressively decays back to 1.0, 4) Beyond the cutoff radius for the low-pass filter, the filter is multiplied by another Gaussian and decays to 0. The MTF curve to be applied should be read in from a file containing values for spatial frequency (in reciprocal pixels) and for the MTF, one pair per line. The program has one built-in curve in which the MTF crosses 0.5 at 0.117/pixel. This curve can be adjusted by scaling its axis, which will make it approximately correct for other situations. To apply only low-pass and high-pass filters, omit the-mtfand-stockoptions; to apply only an inverse filter, omit the-lowpassand other options for general filtering. Similarly, to apply only a low-fre- quency amplifier filter, omit the-mtf,-stock, and other general fil- ter options. If the input file is a real image, then without the "-3d" option the program will take the FFT of each section, apply the filter, take the inverse FFT, and write out the filtered section. With the "-3d" option, it will load the whole file into memory, tapered and padded just as in Taperoutvol, take the 3D FFT, filter, inverse FFT, and write the volume. If the input file is a Fourier transform, it must be a 3D FFT (obtained from "clip fft -3d" or "fftrans -3d"). In this case the program will apply the filter to the transforms in three dimensions and write out a filtered FFT. The program allocates memory dynamically, so it is capable of filtering a rather large volume in 3D. However, it will require 4 bytes of mem- ory per voxel; e.g., 4 GB for a 1 gigavoxel volume. If the program claims that the padded volume is above the memory limit (which will depend on the amount of memory in the computer), or if it fails to allocate the memory, you can chop a volume into pieces, filter them, and reassemble the result. To filter a large image file in 3D this way, simply make a file filterbig.com with one line: $mtffilter -3d <filtering options> INPUTFILE OUTPUTFILE where you insert your filtering options, but INPUTFILE and OUTPUTFILE are exactly as shown, and not the names of your actual input and output files. Then run: chunksetup -p 0 -m 250 filterbig.com input_file output_file where "-p 0" eliminates padding because Mtffilter will take care of padding, "-m 250" specifies the number of megavoxels per chunk, and this time you do put your actual input and output file names. See Chunksetup for details. You can execute the resulting command files with parallel processing (via Processchunks or Etomo) or sequen- tially with: subm filterbig-all Chopping up a volume used to be suggested as a quicker way to filter a large volume, because the time per voxel increases as the log of the volume size, but with current FFT speeds this is much less of an issue.OPTIONSMtffilter uses the PIP package for input (see the manual page for pip). 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 -). Options can be abbreviated to unique letters; the currently valid abbreviations for short names are shown in paren- theses.-input(-inp)OR-InputFileFilenameInput file with images to be filtered-output(-ou)OR-OutputFileFilenameOutput file for filtered images. If this file is omitted, the program will write filtered images back to the input file.-zrange(-z)OR-StartingAndEndingZTwointegersFirst and last Z values in the file to filter. Values are num- bered from 1 and the default is to do all sections. This entry is allowed when doing a 3D FFT on real-space input, but not for FFT input.-mode(-mo)OR-ModeToOutputIntegerThe storage mode of the output file; 0 for byte, 1 for 16-bit signed integer, 6 for 16-bit unsigned integer, 2 for 32-bit floating point, or 12 for 16-bit floating point. The default is the mode of the input file, although the default mode of float- ing point output for MRC files is governed by the value of envi- ronment variable IMOD_WRITE_FLOATS_16BIT. Mode 12 is allowed only if the output format is MRC. This entry is allowed only when writing to a new output file and when the input is not an FFT.-3dfilter(-3)OR-FilterIn3DFilter data in 3D instead of in 2D. The entire volume will be filtered, so it must fit into the allowed amount of memory and-zrangecannot be entered. If the volume will not fit in mem- ory, you can try to use "clip fft -3d" to get an FFT, run Mtf- filter on the 3D FFT, then inverse transform with "clip fft -3d -m mode", where mode is the desired output mode, typically the same as the input. Otherwise you will need to process it in chunks.-1dfilter(-1)OR-OneDimensionalFilterFilter data in 1D (in X direction) instead of in 2D-lowpass(-l)OR-LowPassRadiusSigmaTwofloatsCutoff radius and sigma for a low pass filter that imposes a high-frequency Gaussian roll-off to 0.0. The default is no high-frequency filtering. These entries correspond to the Radius2 and Sigma2 entries to Enhance and other programs; see the Enhance or Filterplot man pages for a full explanation of the effects of changing the sign of the Sigma2 or the Sigma1 and Radius1 parameters entered with the next two options.-highpass(-hi)OR-HighPassSigmaFloatingpointSigma for a high pass filter based on an inverted Gaussian that starts at 0.0 at zero frequency and decays up to 1 with the given sigma. The default is no low-frequency filtering. This entry corresponds to the Sigma1 entry to Enhance and other programs. A negative Sigma1 can be used to get a band-pass fil- ter based on the second derivative of a Gaussian.-radius1(-ra)OR-FilterRadius1FloatingpointCutoff radius for a high-pass filter that is 1.0 at this radius and falls off as a Gaussian to the left of this point with sigma equal to the Sigma2 value entered with -lowpass. This entry corresponds to the Radius1 entry to Enhance and other pro- grams. A negative Radius1 will make the inverted Gaussian invoked by -highpass be zero out to |Radius1|.-mtf(-mt)OR-MtfFileFilenameFile with MTF curve. The format of the file is a series of lines, with a spatial frequency in reciprocal pixels and an MTF value on each line.-stock(-st)OR-StockCurveIntegerThe number of the stock (built-in) MTF curve to use. Since there is only one curve, only an entry of 1 is allowed.-maxinv(-ma)OR-MaximumInverseFloatingpointMaximum value for inverse of MTF. The inverse should always be limited to reduce noise.-invrolloff(-inv)OR-InverseRolloffRadiusSigmaTwofloatsRadius and sigma for gaussian roll-off of inverse to 1.0 (default 0.12 and 0.05)-xscale(-x)OR-XScaleFactorFloatingpointScaling factor for X-axis of MTF curve. Scaling the X axis is probably an adequate way to adapt a curve from one camera or binning to another.-noise(-n)OR-NoisePaddingUse tapered noise based on nearby pixels along the edge of an image to pad an image before filtering. The default is to pad with a simple taper that makes streaks outside the edge of the image. For low-dose images, low-pass filtering can spread these streaks into the image area after cropping and produce artifacts along the edge. This option will avoid this effect, but is probably not suitable for higher-contrast images.-denscale(-den)OR-DensityScaleFactorFloatingpointScaling factor for image intensities after filtering.-rweight(-rw)OR-RWeightedFilterApply an R-weighted filter in the X-dimension, as in back-pro- jection. This option implies-1dfilter. It cannot be used along with an inverse MTF filter. The filter will be scaled to be 1.0 at the cutoff radius specified with the-lowpassoption, if any, or at a frequency of 0.5. This will likely result in a smaller range for the output values, which could lose intensity resolution by making integer values occupy too few gray levels. To overcome this problem, use the-denscaleoption to scale the data up, or change the output mode to floating point with "-mode 2".-fake(-f)OR-FakeSIRTiterationsListofintegerrangesApply a filter to a standard R-weighted back-projection that is equivalent to doing a given number of iterations of SIRT. See the Tilt man page section on SIRT for a description of the filter and the literature reference. If one number is entered, the output file will have the supplied name. If a list of num- bers (which can include ranges) is entered, a file is produced for each entered number, named by appending the number to the entered output name. This filter can be combined with others (sensibly or not). In particular, it could be combined with-rweightand and applied in 1-D to tilt series. It cannot be applied in 3-D, and when applied in 2-D, it is appropriate only when applied to a reconstruction in its original orientation as a stack of X/Z slices. In the latter case, the output here will match the output of Tilt very closely when there is no X-axis tilt or local alignments, and it will diverge increasingly with increasing X-axis tilt. Whether these differences are signifi- cant depends on your application. In any case, applying multi- ple filters would be an efficient way to find the desired number of iterations. Iteration numbers must be under 1000.-pixel(-pi)OR-PixelSizeFloatingpointPixel size in nanometers. A pixel size is needed for dose weighting, deconvolution filtering, and to compute the cutoff radius for an amplifier filter from the phase plate parameters. This entry is needed only if the pixel size in the image file header is incorrect.-volt(-vo)OR-VoltageIntegerMicroscope voltage in kV, used for dose weighting and the decon- volution filter. The default is 300. For dose-weighting, this value must be either 200 or 300; when 200 is entered, the com- puted critical and optimal doses are multiplied by 0.8.DOSEWEIGHTFILTERINGOPTIONSThese options control "dose weighting", which filters out high fre- quencies as a function of the dose already applied to a cryo-specimen.-dtype(-dt)OR-TypeOfDoseFileIntegerThis option both enables dose weighting and indicates what kind of file is being provided with the dose information. Types 1 to 3 are simple text files with a line for each image in the input file containing one or two numbers. These types contain just the dose for each image (type 1), the prior accumulated dose followed by the image dose (type 2), or the prior dose followed by the cumulative dose at the end of that image (type 3). Type 4 indicates information in the autodoc format: either the ".mdoc" file produced by SerialEM, or an HDF file that has incorporated that information into its metadata. In the latter case, no filename would be entered with the-dfileoption. Whichever form is used, the information in the file must be in the same order as the images in the tilt series file. An origi- nal .mdoc file from SerialEM will not work if the input image file has been reordered so that the part of the series acquired first is at the end of the stack.-dfile(-dfil)OR-DoseWeightingFileFilenameName of file with dose information for dose weighting, or a short entry indicating how to derive the file name from the image input file name. This entry is required if-dtypeis entered, unless a 4 is entered and the input file is an HDF file with the equivalent metadata as in an .mdoc file. Two kinds of entries can be used to derive the .mdoc name from the input name. 1) An entry with just an extension and starting with a period (like ".mrc") indicates to replace the extension of the input file with that entry and append ".mdoc". For example, if the input is "setname.ali", an entry of ".mrc" will make it use the filename "setname.mrc.mdoc". 2) An entry starting with "_" and consisting of both a suffix and an extension will make it strip out the suffix from the input name (provided it is just before the extension), substitute the extension in the entry for the one in the input name, and append ".mdoc". For example, if the input is "setname_ali.mrc", an entry of "_ali.mrc" will make it seek "setname.mrc.mdoc" and an entry of "_ali.st" will make it look for "setname.st.mdoc".-dfixed(-dfix)OR-FixedImageDoseFloatingpointFixed dose for each image of the input file, in electrons/square Angstrom. This option cannot be entered with-dtype.-initial(-ini)OR-InitialDoseFloatingpointDose applied before any of the images in the input file were taken; this value will be added to all the prior dose values, however they were obtained.-bidir(-b)OR-BidirectionalNumViewsIntegerNumber of views in the first part of a bidirectional tilt series file, where the order of images in the input file is inverted from their order of acquisition. This entry is essential for a bidirectional series if a fixed dose is entered with-dfixedor if just a dose for each image (dose file type 1) is entered. The program assumes that the part of the tilt series acquired first occurs at the beginning of the stack. If this is not the case, you must enter the-reversedoption. The-bidiroption is ignored for dose file types 2 and 3. It would be used for type 4 if there are neither PriorRecordDose nor DateTime entries; the latter allow the initial order of images to be deduced when there are no PriorRecordDose entries. Intact .mdoc files from SerialEM always have DateTime entries; PriorRecordDose was added in 2017.-reversed(-re)OR-ReversedBidirectionalA bidirectional series is stacked with the part acquired first at the end, and the second part at the beginning. This entry is needed to make the-bidiroption perform properly in the cases described there where it is needed.-optimal(-op)OR-OptimalDoseScalingFloatingpointFactor by which to scale the computed optimal and critical doses that determine how much to attenuate a spatial frequency for a particular dose. Enter a factor greater than or less than 1. to indicate that the specimen is more or less resistant to damage than the equations indicate. Another use for this entry would be to adjust the critical dose for a voltage other than 200 and 300 kV.-critical(-cr)OR-CriticalDoseFactorsThreefloatsReplacement factors a, b, and c in the equation critical_dose = a * k^b + c where k is frequency in reciprocal Angstroms. The default fac- tors are directly from Grant and Grigorieff and the unblur pro- gram. Enter 0 for any of the factors to use the default for that factor.-verbose(-ve)OR-VerboseOutputIntegerEnter a 1 to output the cumulative and image doses used and the filter functions applied for dose weighting.DECONVOLUTIONFILTEROPTIONSThese options control a deconvolution filter like the one implemented in Warp and IsoNet. The filter compensates for a CTF less than 1 at lower frequencies and attenuates higher frequencies.-deconv(-dec)OR-DeconvolutionStrengthFloatingpointOverall strength of the deconvolution, which controls the degree to which low frequencies are boosted. Values around 0.25 to 1 may be useful. The bigger the value, the more low frequencies are accentuated. This value is treated as it is in Warp; values used in IsoNet need to be divided by 3 to have the same effect here.-snr(-sn)OR-SNRFalloffFloatingpointFalloff rate with frequency of the signal-to-noise ratio assumed for computing the deconvolution filter; higher values attenuate high frequencies more. Some users suggest keeping this value comparable to the-deconventry. The default is 0.7, the value in IsoNet.-dchigh(-dch)OR-HighPassNyquistFloatingpointCutoff frequency of a high-pass filter to prevent excessive boosting of the lowest frequencies, where the CTF is low. For consistency with Warp and IsoNet, the value is expressed as the fraction of Nyquist, and is thus twice the usual frequency in reciprocal pixels. The default is 0.02 as in IsoNet.-defocus(-def)OR-DefocusInMicronsFloatingpointThe defocus in microns (underfocus positive). This entry is required for computing the CTF curve; there is no default.-dcphase(-dcp)OR-PhaseShiftFloatingpointPhase shift in degrees (default 0)-csOR-SphericalAberrationFloatingpointSpherical aberration (Cs) in mm (default 2.7)PHASEPLATEFILTEROPTIONSThese options control an amplifier filter used to correct artifacts in images obtained with phase plates having a hole.-amplifier(-a)OR-AmplifierFactorAndPowerTwofloatsAmplification factor and exponent for low-frequency amplifier filter. Either -cutoff or -phase must also be entered.-cutoff(-cu)OR-CutoffForAmplifierFloatingpointCutoff radius in reciprocal pixels for amplifier filter. This option cannot be entered together with -phase.-phase(-ph)OR-PhasePlateParametersThreefloatsParameters of phase plate imaging, used to compute a nominal cutoff radius for the low-frequency amplifier filter. Enter the phase plate diameter in nanometers, the voltage in kilovolts, and the objective lens focal length in millimeters.-param(-pa)OR-ParameterFileParameterfileRead parameter entries from file-help(-he)OR-usagePrint help output-StandardInputRead parameter entries from standard input.HISTORYAdded to package, 3/30/04 Added ability to operate on 3D FFT, 6/19/04 Added ability to take filter real volume in 3D, 5/20/08BUGSEmail bug reports to mast at colorado dot edu. IMOD 4.12.47 mtffilter(1)