Use spike-in controls for absolute quantitation.
For most biological applications, relative quantitation should suffice. Sometimes, though, absolute numbers matter. The solution: spike-in controls. Add a known quantity of an isotopically labeled form of the desired peptide (or close surrogate), and use its abundance to correlate peak height with molecular abundance. This peptide will behave identically with the peptides in your samples during sample preparation and ionization, but it can be distinguished in the instrument because of the slight difference in size.
Invest in a good instrument. Isotopic-labeling approaches rely on a mass spec's ability to distinguish subtle differences in molecular weight — the difference between 16O and 18O, for instance. In theory, most mass specs can handle that task, but in practice a higher-end instrument will inject a dose of confidence into your work. Look for a spec with mass accuracy and resolution high enough to distinguish between modifications such ...
