Oct 20, 2023

Public workspaceProteomic Analysis of Human Whole Lung Tissue Using Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) Acquisition by Data-Independent Acquisition (DIA) on an Orbitrap Eclipse Tribrid Mass Spectrometer

DOI
https://dx.doi.org/10.17504/protocols.io.x54v9p3bpg3e/v1
  • J P Rose1,
  • M Watson1,
  • B Schilling1,
  • Joanna Bons1
  • 1Buck Institute for Research on Aging
  • Metabolomics Protocols & Workflows
  • Cellular Senescence Network (SenNet) Method Development Community
M A Watson
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DOI: https://dx.doi.org/10.17504/protocols.io.x54v9p3bpg3e/v1
Protocol Citation: J P Rose, M Watson, B Schilling, Joanna Bons 2023. Proteomic Analysis of Human Whole Lung Tissue Using Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) Acquisition by Data-Independent Acquisition (DIA) on an Orbitrap Eclipse Tribrid Mass Spectrometer. protocols.io https://dx.doi.org/10.17504/protocols.io.x54v9p3bpg3e/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: October 20, 2023
Last Modified: October 20, 2023
Protocol Integer ID: 89694
Keywords: proteolytic peptide measurement, proteomic analysis of human whole lung tissue, proteomic analysis, reconstituted peptide elution, reconstituted peptide, tandem mass spectrometry, protein lysate, protein identification, human whole lung tissue, using liquid chromatography, protein, liquid chromatography, orbitrap eclipse tribrid mass spectrometer, orbitrap eclipse tribrid mass spectrometer frozen
Abstract
Frozen human whole lung tissues were homogenized using a urea-based buffer and bead mill homogenizer to isolate protein. The protein lysate was subjected to tryptic digestion using S-trap Spin columns. The reconstituted peptide elution was desalted with C18 hydrophilic-lipophilic balance (HLB) cartridges. The final reconstituted peptides were diluted with 2% ACN and 0.1% FA. Proteolytic peptide measurement was completed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) acquisition by Data-Independent Acquisition (DIA) on an Orbitrap Eclipse Tribrid mass spectrometer for peptide/protein identification and quantification.
Protocol references