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Deep Single-Shot NanoLC-MS Proteome Profiling with a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS
[Image: see text] This study demonstrates how the latest ultrahigh-performance liquid chromatography (UHPLC) technology can be combined with high-resolution accurate-mass (HRAM) mass spectrometry (MS) and long columns packed with fully porous particles to improve bottom-up proteomics analysis with n...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Chemical Society
2022
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9552226/ https://www.ncbi.nlm.nih.gov/pubmed/36068014 http://dx.doi.org/10.1021/acs.jproteome.2c00270 |
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author | Zheng, Runsheng Stejskal, Karel Pynn, Christopher Mechtler, Karl Boychenko, Alexander |
author_facet | Zheng, Runsheng Stejskal, Karel Pynn, Christopher Mechtler, Karl Boychenko, Alexander |
author_sort | Zheng, Runsheng |
collection | PubMed |
description | [Image: see text] This study demonstrates how the latest ultrahigh-performance liquid chromatography (UHPLC) technology can be combined with high-resolution accurate-mass (HRAM) mass spectrometry (MS) and long columns packed with fully porous particles to improve bottom-up proteomics analysis with nanoflow liquid chromatography–mass spectrometry (nanoLC-MS) methods. The increased back pressures from the UHPLC system enabled the use of 75 μm I.D. × 75 cm columns packed with 2 μm particles at a typical 300 nL/min flow rate as well as elevated and reduced flow rates. The constant pressure pump operation at 1500 bar reduced sample loading and column washing/equilibration stages and overall overhead time, which maximizes MS utilization time. The versatility of flow rate optimization to balance the sensitivity, throughput with sample loading amount, and capability of using longer gradients contributes to a greater number of peptide and protein identifications for single-shot bottom-up proteomics experiments. The routine proteome profiling and precise quantification of >7000 proteins with single-shot nanoLC-MS analysis open possibilities for large-scale discovery studies with a deep dive into the protein level alterations. Data are available via ProteomeXchange with identifier PXD035665. |
format | Online Article Text |
id | pubmed-9552226 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | American Chemical Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-95522262022-10-12 Deep Single-Shot NanoLC-MS Proteome Profiling with a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS Zheng, Runsheng Stejskal, Karel Pynn, Christopher Mechtler, Karl Boychenko, Alexander J Proteome Res [Image: see text] This study demonstrates how the latest ultrahigh-performance liquid chromatography (UHPLC) technology can be combined with high-resolution accurate-mass (HRAM) mass spectrometry (MS) and long columns packed with fully porous particles to improve bottom-up proteomics analysis with nanoflow liquid chromatography–mass spectrometry (nanoLC-MS) methods. The increased back pressures from the UHPLC system enabled the use of 75 μm I.D. × 75 cm columns packed with 2 μm particles at a typical 300 nL/min flow rate as well as elevated and reduced flow rates. The constant pressure pump operation at 1500 bar reduced sample loading and column washing/equilibration stages and overall overhead time, which maximizes MS utilization time. The versatility of flow rate optimization to balance the sensitivity, throughput with sample loading amount, and capability of using longer gradients contributes to a greater number of peptide and protein identifications for single-shot bottom-up proteomics experiments. The routine proteome profiling and precise quantification of >7000 proteins with single-shot nanoLC-MS analysis open possibilities for large-scale discovery studies with a deep dive into the protein level alterations. Data are available via ProteomeXchange with identifier PXD035665. American Chemical Society 2022-09-06 2022-10-07 /pmc/articles/PMC9552226/ /pubmed/36068014 http://dx.doi.org/10.1021/acs.jproteome.2c00270 Text en © 2022 The Authors. Published by American Chemical Society https://creativecommons.org/licenses/by-nc-nd/4.0/Permits non-commercial access and re-use, provided that author attribution and integrity are maintained; but does not permit creation of adaptations or other derivative works (https://creativecommons.org/licenses/by-nc-nd/4.0/). |
spellingShingle | Zheng, Runsheng Stejskal, Karel Pynn, Christopher Mechtler, Karl Boychenko, Alexander Deep Single-Shot NanoLC-MS Proteome Profiling with a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS |
title | Deep Single-Shot
NanoLC-MS Proteome Profiling with
a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS |
title_full | Deep Single-Shot
NanoLC-MS Proteome Profiling with
a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS |
title_fullStr | Deep Single-Shot
NanoLC-MS Proteome Profiling with
a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS |
title_full_unstemmed | Deep Single-Shot
NanoLC-MS Proteome Profiling with
a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS |
title_short | Deep Single-Shot
NanoLC-MS Proteome Profiling with
a 1500 Bar UHPLC System, Long Fully Porous Columns, and HRAM MS |
title_sort | deep single-shot
nanolc-ms proteome profiling with
a 1500 bar uhplc system, long fully porous columns, and hram ms |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9552226/ https://www.ncbi.nlm.nih.gov/pubmed/36068014 http://dx.doi.org/10.1021/acs.jproteome.2c00270 |
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