A comprehensive pipeline for translational top-down proteomics from a single blood draw

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Date

2016-11

Journal Title

Journal ISSN

Volume Title

Publisher

Nature Protocols

Abstract

Top-down proteomics (TDP) by mass spectrometry (MS) has become increasingly popular in translational spheres as researchers discern the value of proteoform-resolved measurements. The advent of robust prefractionation strategies for intact proteoforms and the emerging availability of high-resolution benchtop mass spectrometers have catalyzed the emergence of high-throughput TDP for the analysis of biological and clinical samples. Here, we provide a comprehensive protocol for TDP of a sample type often investigated in translational research: human peripheral blood mononuclear cells (PBMC). The workflow described herein comprises sample collection, cell lysis, proteoform prefractionation, instrument setup, liquid chromatography-tandem MS (LC-MS/MS) analysis, and data processing, for a minimum completion time of 48 hours. Importantly, this workflow is compatible with multiple PBMC processing strategies and existing biorepository practices, can be used for both small-scale targeted proteoform studies or large-scale multi-sample discovery studies, and has been successfully applied in quantitative contexts for biomarker discovery.

Description

Keywords

Top-down proteomics, Translational proteomics, Mass spectrometry, Proteoform, Proteomics

Citation

Forthcoming

DOI

Forthcoming

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The work (as defined below) is provided under the terms of this creative commons public license ("ccpl" or "license"). The work is protected by copyright and/or other applicable law. Any use of the work other than as authorized under this license or copyright law is prohibited. By exercising any rights to the work provided here, you accept and agree to be bound by the terms of this license. To the extent this license may be considered to be a contract, the licensor grants you the rights contained here in consideration of your acceptance of such terms and conditions.

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