Patent ID: 7655433
Filing Date: 2010-02-02
Classification: G01N

Abstract:
1. A method of identifying and quantifying polypeptides in a sample, comprising the steps of: (a) cleaving a polypeptide sample to generate sample peptides; (b) labeling the sample peptides with a first isotope tag to provide a sample of a first isotopically-labeled peptide; (c) chemically synthesizing a plurality of peptide standards of known sequences, wherein the chemically synthesized peptide standards of known sequences comprise sequences of peptides generated by the same cleaving as in step (a); (d) labeling the chemically synthesized peptide standards with a second isotope tag, wherein the second isotope tag labels the chemically synthesized peptide standards with an isotopically distinct version of the first isotope tag, thereby generating a plurality of isotopically-labeled chemically synthesized peptide standards; (e) adding the plurality of the isotopically-labeled chemically synthesized peptide standards of known sequences to the isotopically-labeled sample peptides, thereby generating a mixture of isotopically-labeled sample peptides and isotopically-labeled chemically synthesized peptide standards; (f) resolving the mixture of labeled sample peptides and labeled chemically synthesized peptide standards into a plurality of resolved fractions; (g) analyzing the resolved fractions by mass spectrometry; (h) identifying an isotopically-labeled sample peptide in an analyzed fraction from step (g); and (i) determining the amount of the identified isotopically-labeled sample peptide in the analyzed fraction by comparing the amount of the identified isotopically-labeled sample peptide to the amount of the isotopically-labeled chemically synthesized peptide standard in the analyzed fraction.