Label-Free Quantitation of Endogenous Peptides.

Methods Mol Biol

Department of Chemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.

Published: April 2024

Liquid chromatography-mass spectrometry (LC-MS)-based peptidomics methods allow for the detection and identification of many peptides in a complex biological mixture in an untargeted manner. Quantitative peptidomics approaches allow for comparisons of peptide abundance between different samples, allowing one to draw conclusions about peptide differences as a function of experimental treatment or physiology. While stable isotope labeling is a powerful approach for quantitative proteomics and peptidomics, advances in mass spectrometry instrumentation and analysis tools have allowed label-free methods to gain popularity in recent years. In a general label-free quantitative peptidomics experiment, peak intensity information for each peptide is compared across multiple LC-MS runs. Here, we outline a general approach for label-free quantitative peptidomics experiments, including steps for sample preparation, LC-MS data acquisition, data processing, and statistical analysis. Special attention is paid to address run-to-run variability, which can lead to several major problems in label-free experiments. Overall, our method provides researchers with a framework for the development of their own quantitative peptidomics workflows applicable to quantitation of peptides from a wide variety of different biological sources.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC11027169PMC
http://dx.doi.org/10.1007/978-1-0716-3646-6_7DOI Listing

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