A method for label-free, differential top-down proteomics

Ioanna Ntai, Timothy K. Toby, Richard D. Leduc, Neil L. Kelleher

Research output: Chapter in Book/Report/Conference proceedingChapter

20 Scopus citations

Abstract

Biomarker discovery in the translational research has heavily relied on labeled and label-free quantitative bottom-up proteomics. Here, we describe a new approach to biomarker studies that utilizes high throughput top-down proteomics and is the first to offer whole protein characterization and relative quantitation within the same experiment. Using yeast as a model, we report procedures for a label-free approach to quantify the relative abundance of intact proteins ranging from 0 to 30 kDa in two different states. In this chapter, we describe the integrated methodology for the large-scale profiling and quantitation of the intact proteome by liquid chromatography-mass spectrometry (LC-MS) without the need for metabolic or chemical labeling. This recent advance for quantitative top-down proteomics is best implemented with a robust and highly controlled sample preparation workflow before data acquisition on a high-resolution mass spectrometer, and the application of a hierarchical linear statistical model to account for the multiple levels of variance contained in quantitative proteomic comparisons of samples for basic and clinical research.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc
Pages121-133
Number of pages13
DOIs
StatePublished - Feb 1 2016

Publication series

NameMethods in Molecular Biology
Volume1410
ISSN (Print)1064-3745

Keywords

  • Differential expression
  • Label-free quantitation
  • Proteoform
  • Quantitative mass spectrometry
  • Top-down proteomics
  • Top-down quantitation

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Ntai, I., Toby, T. K., Leduc, R. D., & Kelleher, N. L. (2016). A method for label-free, differential top-down proteomics. In Methods in Molecular Biology (pp. 121-133). (Methods in Molecular Biology; Vol. 1410). Humana Press Inc. https://doi.org/10.1007/978-1-4939-3524-6_8