Label-free peptide quantification (LFQ) typically involves summing MS1 isotope signal intensities across the chromatographic elution window to generate an extracted ion chromatogram (XIC). The XIC is then integrated to calculate the area under the curve, which is used to compare relative peptide abundances.
For post-translational modifications (PTMs) that produce only small mass shifts between modified and unmodified peptides, constructing the XIC from a single isotope is generally sufficient for accurate relative quantification. However, when the mass difference is large enough to alter the isotopic distribution, reliance on a single isotope becomes unreliable.
The Byos PTM report template includes a theoretical all-isotope XIC calculation, termed Total XIC AUC Averagine, which extrapolates the total XIC area from the most intense isotope using the averagine-based theoretical isotopic distribution.
This quantitation strategy is better suited for comparing modified peptides, such as glycosylated species, to their non-glycosylated counterparts, which may be inaccurately represented when single-isotope approaches are used.
To enable this feature from with in the PTM report, select the fourth tab (% Mod (Multi Mods Per Peptide):
Then select Total XIC AUC Averagine from the dropdown menu: