This method is used when the Ratio Calculation Parameter in the Precursor Ions Quantifier and the Reporter Ions Quantifier is set to Summed Abundance Based. When the study design is non-nested, the protein ratio is calculated by dividing the summed abundance values described in Calculating group abundances for the two samples chosen for that ratio. The peptide group ratio is also calculated by dividing the summed abundance values for that specific peptide.
For precursor ion quantification, the peptide group ratios are calculated slightly differently because the various isotopic forms consist of different consensus features. In this case, the peptide group abundances and the ratio are shown for only one of the isotopic forms used for the ratio when more than one of the isotopic forms has a confidently identified PSM.
In this example, the highlighted row shows that the heavy-labeled form of the peptide includes a ratio column and grouped abundance values for both the light and heavy forms of the peptide, while the light form does not show a ratio nor abundance values. For protein quantification, the ratios are calculated as a sum of the individual peptide group abundances as just described.
For a nested study design with biological replicates, the application calculates the peptide group and protein ratios differently. In this case, ratios for the summed abundances for each biological replicate are calculated separately, and the geometric median of the resulting ratios is used as the peptide group and protein ratios. The following figure shows an example calculation of protein ratios for non-nested and nested designs.
Thermo Fisher Scientific recommends that you use the summed abundance approach for reporter ion quantification, such as TMT where there are few missing values. For precursor ion quantification and label-free quantification, use the pairwise ratio approach described in Calculating ratios using the Pairwise Ratio approach.
The Maximum Allowed Fold Change parameter defines the maximum fold change that the node calculates. All ratios are between 1/fold change and fold change. When the numerator or the denominator is 0, the ratio is the fold change when the denominator is 0 or 1/fold change when the numerator is 0.