Mass Identity

What it determines

The sample is separated by liquid chromatography and introduced into the mass spectrometer, where the eluting molecules are ionized. The instrument measures mass-to-charge ratios. Peptides generally carry several charges at once, and the observed charge series is deconvoluted to a single molecular mass.

That observed mass is compared with the mass calculated from the stated sequence. In the second stage, selected precursor ions are fragmented and the resulting fragment masses are mapped back onto the backbone, which confirms identity at the sequence level. Agreement is judged against the tolerance established for the method.

Practical notes

• The expected sequence and any modifications such as amidation, acetylation, or cyclization are supplied with the sample. The comparison is only as good as the sequence given.
• Salt form and adducts shift the observed ions. Sodium and potassium adducts are common and are accounted for during interpretation.
• Material submitted in a heavily salted buffer benefits from a note at intake so the chromatography can be set up for it.