DIA label-free proteomic analysis of murine bone-marrow-derived macrophages

Christa P. Baker; Iain R. Phair; Alejandro J. Brenes; Abdelmadjid Atrih; Dylan G. Ryan; Roland Bruderer; Albena T. Dinkova-Kostova; Douglas J. Lamont; J. Simon C. Arthur; Andrew J. M. Howden

doi:10.1016/j.xpro.2022.101725

DIA label-free proteomic analysis of murine bone-marrow-derived macrophages

Christa P. Baker, Iain R. Phair, Alejandro J. Brenes, Abdelmadjid Atrih, Dylan G. Ryan, Roland Bruderer, Albena T. Dinkova-Kostova, Douglas J. Lamont, J. Simon C. Arthur (Lead / Corresponding author), Andrew J. M. Howden (Lead / Corresponding author)

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Abstract

Here, we describe an optimized protocol to analyze murine bone-marrow-derived macrophages using label-free data-independent acquisition (DIA) proteomics. We provide a complete step-by-step protocol describing sample preparation utilizing the S-Trap approach for on-column digestion and peptide purification. We then detail mass spectrometry data acquisition and approaches for data analysis. Single-shot DIA protocols achieve comparable proteomic depth with data-dependent MS approaches without the need for fractionation. This allows for better scaling for large sample numbers with high inter-experimental reproducibility. For complete details on the use and execution of this protocol, please refer to Ryan et al. (2022).

Original language	English
Article number	101725
Number of pages	17
Journal	STAR Protocols
Volume	3
Issue number	4
Early online date	26 Sept 2022
DOIs	https://doi.org/10.1016/j.xpro.2022.101725
Publication status	Published - 16 Dec 2022

Keywords

Bioinformatics
Immunology
Mass spectrometry
Protein biochemistry
Proteomics

ASJC Scopus subject areas

General Neuroscience
General Biochemistry,Genetics and Molecular Biology
General Immunology and Microbiology

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10.1016/j.xpro.2022.101725Licence: CC BY

Final published versionFinal published version, 2.42 MBLicence: CC BY

Cite this

@article{158722148013409c9154c176df57aae0,

title = "DIA label-free proteomic analysis of murine bone-marrow-derived macrophages",

abstract = "Here, we describe an optimized protocol to analyze murine bone-marrow-derived macrophages using label-free data-independent acquisition (DIA) proteomics. We provide a complete step-by-step protocol describing sample preparation utilizing the S-Trap approach for on-column digestion and peptide purification. We then detail mass spectrometry data acquisition and approaches for data analysis. Single-shot DIA protocols achieve comparable proteomic depth with data-dependent MS approaches without the need for fractionation. This allows for better scaling for large sample numbers with high inter-experimental reproducibility. For complete details on the use and execution of this protocol, please refer to Ryan et al. (2022).",

keywords = "Bioinformatics, Immunology, Mass spectrometry, Protein biochemistry, Proteomics",

author = "Baker, {Christa P.} and Phair, {Iain R.} and Brenes, {Alejandro J.} and Abdelmadjid Atrih and Ryan, {Dylan G.} and Roland Bruderer and Dinkova-Kostova, {Albena T.} and Lamont, {Douglas J.} and Arthur, {J. Simon C.} and Howden, {Andrew J. M.}",

note = "Funding Information: This work was funded by the Wellcome Trust ( 102132/B/13/Z ) and Cancer Research UK ( C20953/A18644 ). Publisher Copyright: {\textcopyright} 2022 The Author(s)",

year = "2022",

month = dec,

day = "16",

doi = "10.1016/j.xpro.2022.101725",

language = "English",

volume = "3",

journal = "STAR Protocols",

issn = "2666-1667",

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AU - Baker, Christa P.

AU - Phair, Iain R.

AU - Brenes, Alejandro J.

AU - Atrih, Abdelmadjid

AU - Ryan, Dylan G.

AU - Bruderer, Roland

AU - Dinkova-Kostova, Albena T.

AU - Lamont, Douglas J.

AU - Arthur, J. Simon C.

AU - Howden, Andrew J. M.

PY - 2022/12/16

Y1 - 2022/12/16

N2 - Here, we describe an optimized protocol to analyze murine bone-marrow-derived macrophages using label-free data-independent acquisition (DIA) proteomics. We provide a complete step-by-step protocol describing sample preparation utilizing the S-Trap approach for on-column digestion and peptide purification. We then detail mass spectrometry data acquisition and approaches for data analysis. Single-shot DIA protocols achieve comparable proteomic depth with data-dependent MS approaches without the need for fractionation. This allows for better scaling for large sample numbers with high inter-experimental reproducibility. For complete details on the use and execution of this protocol, please refer to Ryan et al. (2022).

AB - Here, we describe an optimized protocol to analyze murine bone-marrow-derived macrophages using label-free data-independent acquisition (DIA) proteomics. We provide a complete step-by-step protocol describing sample preparation utilizing the S-Trap approach for on-column digestion and peptide purification. We then detail mass spectrometry data acquisition and approaches for data analysis. Single-shot DIA protocols achieve comparable proteomic depth with data-dependent MS approaches without the need for fractionation. This allows for better scaling for large sample numbers with high inter-experimental reproducibility. For complete details on the use and execution of this protocol, please refer to Ryan et al. (2022).

KW - Bioinformatics

KW - Immunology

KW - Mass spectrometry

KW - Protein biochemistry

KW - Proteomics

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DO - 10.1016/j.xpro.2022.101725

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JF - STAR Protocols

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M1 - 101725

ER -

DIA label-free proteomic analysis of murine bone-marrow-derived macrophages

Abstract

Keywords

ASJC Scopus subject areas

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Molecular and Cellular Biology 4 Year PhD Studentships

The Role of the Keap1/Nrf2 Pathway in Tumour Metabolic Adaptation (Joint with University of Cambridge and University College London)

Investigating the murine bone marrow derived macrophage response to Candida albicans infection

Cite this

DIA label-free proteomic analysis of murine bone-marrow-derived macrophages

Abstract

Keywords

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Projects

Molecular and Cellular Biology 4 Year PhD Studentships

The Role of the Keap1/Nrf2 Pathway in Tumour Metabolic Adaptation (Joint with University of Cambridge and University College London)

Student theses

Investigating the murine bone marrow derived macrophage response to Candida albicans infection

Cite this