For precursor ion quantification, the Quantification Spectrum chart displays a quantification spectrum for each peptide. It also displays the different abundances of the identified Light, Medium, and Heavy isotopic peak patterns used to quantify a peptide. The abundances are measured by calculating the area of the extracted ion chromatogram of each isotope of a pattern. The chart highlights the corresponding isotope pattern peaks and labels them with the quantification channel names. The chart also includes any peaks that are not part of an isotope pattern.

Procedure

  1. Select the PSMs tab in the result file.
  2. Select the row of the PSM that interests you.
  3. Choose View > Quantification Spectrum, or select Quantification Spectrum .
  4. Quantification Spectrum chart for precursor ion quantification
  5. The x axis of the chart displays the mass-to-charge ratio of the isotopes, and the y axis displays the area of the extracted ion chromatogram for the isotopes. Filled blue circles mark the isotope pattern peaks for the different isotopic forms of the identified peptide. Only the most abundant peak is used for quantification. Other isotope distributions that coelute are labeled with diamonds in different colors, with peaks from the same isotope clusters having the same color of diamond.
  6. The Quantification Spectrum chart can also indicate whether an expected quantification pattern peak is absent. Regions in pink indicate where a quantification pattern peak was expected but is absent. This ion pattern peak is not used in calculating the quantification values for the different quantification channels.
  7. Expected but absent peak in the Quantification Spectrum chart
  8. Regions in green indicate where a quantification pattern peak was expected but is unsuitable. Pattern peaks might be unsuitable because of the wrong centroid retention time, out-of-delta mass range, wrong intensity, or a peak that has been used by another isotopic pattern. This ion pattern peak is not used in calculating the quantification values for the different quantification channels.
  9. Expected but unsuitable peaks in the Quantification Spectrum chart

The following table shows what the various colors mean on the Quantification Spectrum charts

The meaning of colors in the Quantification Spectrum chart

Color

Meaning

Filled blue circle

Indicates the isotope pattern peaks that are used in calculating the quantification values for the different quantification channels.

Yellow box

Indicates that the pattern includes peaks from indistinguishable channels. This ion pattern peak is not used in calculating the quantification values for the different quantification channels.

Pink bar

Indicates that a quantification pattern peak is expected but is missing. This ion pattern peak is not used in calculating the quantification values for the different quantification channels.

Green bar

Indicates that a quantification pattern peak is present but is unsuitable because of errors in peptide labeling or because of the wrong centroid retention time, the out-of-delta mass range, the wrong intensity, or a peak that has been used by another isotopic pattern. This ion pattern peak is not used in calculating the quantification values for the different quantification channels.