Patent ID: 7244411
Filing Date: 2007-07-17
Classification: C12Q,G01N

Abstract:
1. A method for identification and quantification of one or more proteins in complex mixtures wherein the peptides not containing histidine nor arginine, named here NHNR peptides, are selectively isolated from each protein and a relative concentration of one or more proteins is determined from the ratio between the areas of estimated theoretical spectra for each NHNR peptide labeled with different isotopes in each sample, the method comprising the following steps: a. enzymatically or chemically hydrolyzing the sample or samples of proteins; b. chemically modifying alpha and epsilon amino groups (α and ε-NH2) of every peptide obtained in step (a) with an agent that neither contains, nor generates positively charged groups, nor basic groups, nor possible sites of protonation; c. isolating the NHNR peptides by cation exchange chromatography from the mixture of peptides obtained in step (b); d. identifying proteins by mass spectrometry analysis of the NHNR peptides obtained in step (c); e. differential isotopic labeling of protein samples previously to step (a) or during steps (a) or (b) and immediately mixing at least a portion of the samples; and f. relative quantifying one or more proteins in the mixtures of step (e) from the ratio between the areas of estimated theoretical spectra of the NHNR peptides identified in step (d), as well as from the ratio between the areas of the estimated theoretical spectra of fragments from the NHNR peptides, generated in step (d).