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Assessing LC-MS Data Quality

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Running MScout
The general format for usage for the 64-bit version is
% mscout64 <options> <input MS1 file(s)>

For the 32-bit version, simply replace mscout64 with mscout32.

MScout currently accepts only MS1 files as input. See reference [3] at the bottom of the MScout home page for details. Download MakeMS2 to convert your RAW files from Thermo Scientific instruments into the MS1 format.

An example command is

% mscout64 -rti=0 -rtf=120 -time=0.2 -mz=0.5 -sn=3 -histograms=0 -siglist=1 ./ms1-files/052107-faims-yeast-03-CV-*.ms1


  • rti, rtf: initial retention time & final retention time, between which calculations are performed [default: maximum initial retention time and minimum final retention time, read from the input files]
  • time: time in minutes over which signals should persist in order to be counted as peptide signals [default = 0.2 min]
  • mz: approximate m/z resolution of the instrument that generated the data [default = 0.5]
  • sn: Signal-to-noise ratio threshold for counting signals -- everything over thi s value is examined; [default = 3]
  • order: determines binning for generating intensity and S/N histograms; leave it out [default = 0.01]
  • skipzero: Tells the signal-to-noise algorithm to skip zeros in the data set whe n computing noise level (1 = yes, 0 = no) [default = 1]
  • compare: indicate whether you intend to compare signals from multiple files (1 = yes, 0 = no) [default = 0]
  • centroid: indicate whether your input files contain centroided data (1 = yes, 0 = no) [default = 0]
  • histograms: indicates whether you want Mscout to generate intensity and S/N hist ograms for your data (1 = yes, 0 = no) [default = 1]
  • siglist: indicates whether you want Mscout to generate a list of all detected pe ptide signals (1 = yes, 0 = no) [default = 1]
  • output: the name of the file to which analysis results (e.g. peak capacity & dynamic range info) will be written [default = mscout-results.txt
  • location: indicates whether you want dynamic range calculations to be done at the top of a chromatographic peak (location=0), at the leading edge (location=1), or at the trailing edge (location=2) [default = 0]
  • input MS1 file: the input MS1 file or files
MScout Output

MScout's output is in three parts, two of which are optional:

  1. mscout-results.txt (not optional, but filename can be changed by user). This file contains summary information about the data run (date, input parameters), as well as the following quantities:
    • Peak capacity in the time domain
    • Dynamic range
    • Intensities of most intense and least intense peaks.
    • Number of detected peptide-like peaks.
    • Signal-to-noise of the most-intense and least-intense peaks.
  2. List of signals, with filename SL-<input filename>.txt (optional). This file contains a list of all detected peptide-like signals. Each line of the file contains:
    • m/z of the signal
    • Start, peak, and end points of the signal in the time domain
    • Intensity at the start, peak, and end
    • Signal-to-noise at the start, peak, and end
    • Which additional runs (for FAIMS, CV values) this signal appears in
  3. Histograms (optional). Selecting this option generates intensity and signal-to-noise histograms from the input data.
    • Intensity: filename INT-<input filename>.txt
    • Signal-to-noise: filename SN-<input filename>.txt
  1. Input data files (or symbolic links to input data files) must be in the same directory from which you launch MScout. If this is not the case, MScout will be confused about where to write output files. This is a known issue which will be resolved in a future version.
  2. Results for high-resolution data (such as that obtained from Fourier transform instruments like FT-ICR or Orbitrap instruments) are unpredictable. MScout is intended for use on low-resolution data. For similar information from high-resolution data, see Hardklör.