Q4SRMcmd usage - PNNL-Comp-Mass-Spec/Q4SRM GitHub Wiki

Q4SRMcmd is the command-line version of Q4SRM.

Primary Inputs

  • Thermo .RAW SRM file, mzML file, or folder containing multiple such files (tested on files from TSQ Vantage and TSQ Altis)
    • 64-bit OS: Thermo .RAW files are read using Thermo's RawFileReader library, more information available at Planet Orbitrap
      • A license document for use of this library is included in the source code folder 'Lib', and also will be included in the binary download/installer packages.
    • 32-bit OS: Thermo .RAW files are read using Thermo's MSFileReader library, which must be previously installed
    • NOTE: Currently does not support mzML chromatograms; if creating the mzML file with ProteoWizard/msconvert, specify the '--srmAsSpectra' option
  • If reading mzML file(s): a CSV/TSV file with the following columns:
    • 'Compound Name': Name of the compound, also used to determine heavy compounds by finding "heavy" or "hvy" in the compound name
    • 'Precursor (m/z)': Precursor m/z
    • 'Product (m/z)': Product/transition m/z
    • 'Start Time (min)': Elution window start time
    • 'End Time (min)': Elution window end time
  • Metrics controls
    • (Optional) A TSV file with compound and threshold columns, for custom thresholds for the specified compounds
    • A summed intensity minimum passing value (group of transitions); default is 10000
    • A minimum distance (in minutes) that a 'peak apex' must be from the edges of its elution window (single transition); default is 0.5 (minutes)
    • A maximum distance (in minutes) from the earliest 'peak apex' to the latest 'peak apex' in the compound (group of transitions); default is 0.5 (minutes)
    • A minimum difference of the max intensity point vs. the median of intensity (single transition); default is 10


  • A TSV file with compound and transition information, and values signifying whether the compound/transitions passed or failed a metric
  • An image file (default is PNG format, can also be JPEG, SVG, or PDF) showing a scatterplot of compounds with marks showing if they failed certain metrics, with axes of 'max intensity time' and 'max intensity'.
  • A PDF that includes the scatterplot mentioned above, and also showing the chromatograms of each transition in each group.
  • HeavySummary.tsv, which is a short summary file listing total heavy peptides and the number that passed the metrics for each dataset

Syntax & Usage

Q4SRMcmd.exe -raw file.raw [other options]

Full usage information:

  -?, -help           Show this help screen

  -raw                Required. Path to .raw file, or directory containing .raw

  -recurse            If raw file path is a folder, whether to find .raw files
                      in subfolders (Default: False)

  -filter             If raw file path is a folder, a file filter string
                      (supports '*' wildcard) (Default: *.raw)

  -method             For mzML files, the method file path

  -t                  Peak area threshold for a compound to be considered
                      "passing" (Default: 10000, Min: 0)

  -m                  Time (in minutes) that a peak must be away from the edge
                      of the target window for it to be considered "passing"
                      (Default: 0.5, Min: 0)

  -ec                 Threshold (in minutes) for elution concurrence of the
                      transition peaks for the same compound; smaller is
                      stricter. (Default: 0.5, Min: 0)

  -sn                 Threshold for the Signal-to-Noise heuristic; larger is
                      stricter; value is calculated as max intensity / median
                      intensity (Default: 10, Min: 1)

  -tpc                A TSV file with compound and threshold columns, for
                      custom thresholds for the specified compounds
                      (Default: "")

  -out                Folder where the result files should be written (default:
                      written to same folder as .raw)

  -threads            Maximum number of threads to use (files processed
                      simultaneously), '0' for automatic (Default: 0, Min: 0)

  -ow, -overwrite     If specified, all files will be processed, even if
                      existing output was created with the same settings.
                      (Default: False)

  -outThresholds      If specified, creates a per-compound thresholds file for
                      all compounds that pass the minimum threshold (averaged
                      across processed files) (Default: False)

  -outThresholdsPct   The percentage of the averaged total intensity that
                      should be output as the threshold (Default: 0.5,
                      Min: 0.01, Max: 0.99)

  -outThresholdsPath  The path where the output threshold file should be

  -summaryPath        Path where processing summary tsv should be written.
                      Default is [out folder/raw file folder]\HeavySummary.tsv

  -img                Format for the saved total intensity vs. time plot
                      (Default: PNG (or 0))
                      Possible values are:
                        0 or 'PNG'
                        1 or 'PDF'
                        2 or 'SVG'
                        3 or 'JPEG'
                        -1 or 'NoImageExport': Don't save an image

  -all                If specified, metrics are computed for all compounds in
                      the file (not just heavy compounds) (Default: False)