Quantitative analysis of complex peptide mixtures using FTMS and differential mass spectrometry

Abstract

Label-free LC-MS profiling is a powerful quantitative proteomic method to study relative peptide abundances between two or more biological samples. Here we demonstrate the use of a previously described comparative LC-MS method, differential mass spectrometry (dMS), to analyze high-resolution Fourier transform mass spectrometry (FTMS) data for detection and quantification of known peptide differences between two sets of complex mixtures. Six standard peptides were spiked into a processed plasma background at fixed ratios from 1.25:1 to 4:1 to make two sets of samples. The resulting mixtures were analyzed by microcapillary LC-FTMS and dMS. dMS successfully identified five out of the six peptides as statistically significant differences (p Յ 0.005). In this experiment, the smallest fold change reliably detected by our method was 1.5:1, and the errors of estimated ratios of concentrations were less than 20% for peptides spiked at 1.5:1 to 4:1. We conclude that LC-FTMS coupled with dMS is a useful label-free quantitative MS method that can be used to detect subtle yet statistically significant peptide differences in complex protein mixtures, including plasma samples. (J Am Soc Mass Spectrom 2007, 18, 226 -233)

Proteomic Profiling of a Layered Tissue Reveals Unique Glycolytic Specializations of Photoreceptor Cells

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Diversity of human skeletal muscle in health and disease: Contribution of proteomics

“…Subjects were monitored at 5 different times, before and after the end of exercise. Plasma underwent glycopeptide enrichment, followed by LC MS/MS [98][99][100]. Potential protein markers with at least 2 distinct peptide components increasing during exercise as a function of work load and duration, were selected.…”

Quantitative analysis of intact apolipoproteins in human HDL by top-down differential mass spectrometry

“…Differential mass spectrometry has been successfully applied for the relative quantitation of complex peptide mixtures (17,18). To adapt dMS to the analysis of intact protein mixtures, two primary requirements to retain are a reproducible biochemical sample processing method and a robust LC-MS analysis.…”

“…To identify proteins of interest (as defined by m∕z ratio, intensity and retention time) we used dMS, a label-free LC-MS method for finding ions that exhibit a statistically significant difference in abundance in complex mixtures of peptides or proteins as previously described (8,17,18). Expression profiles generated from the highresolution LC-MS data above were analyzed using the Rosetta Elucidator™ system (version 3.2) (40).…”

“…Differential mass spectrometry (dMS) is a general proteomics workflow that provides relative quantitation from full-scan mass spectrometry data (17)(18)(19). Here, we set out to establish topdown dMS capabilities that combine the advantages of intact protein analysis with quantitation from full spectrum mass spectrometry data (Fig.…”