Conventional-Flow Liquid Chromatography-Mass Spectrometry for Exploratory Bottom-Up Proteomic Analyses
The result's identifiers
Result code in IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216208%3A11160%2F18%3A10375502" target="_blank" >RIV/00216208:11160/18:10375502 - isvavai.cz</a>
Alternative codes found
RIV/00179906:_____/18:10375502 RIV/60162694:G44__/18:43889525
Result on the web
<a href="http://pubs.acs.org/doi/10.1021/acs.analchem.8b00525" target="_blank" >http://pubs.acs.org/doi/10.1021/acs.analchem.8b00525</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.1021/acs.analchem.8b00525" target="_blank" >10.1021/acs.analchem.8b00525</a>
Alternative languages
Result language
angličtina
Original language name
Conventional-Flow Liquid Chromatography-Mass Spectrometry for Exploratory Bottom-Up Proteomic Analyses
Original language description
Due to its sensitivity and productivity, bottom-up proteomics based on liquid chromatography-mass spectrometry (LC-MS) has become the core approach in the field. The de facto standard LC-MS platform for proteomics operates at sub-mu L/min flow rates, and nanospray is required for efficiently introducing peptides into a mass spectrometer. Although this is almost a "dogma", this view is being reconsidered in light of developments in highly efficient chromatographic columns, and especially with the introduction of exceptionally sensitive MS instruments. Although conventional-flow LC-MS platforms have recently penetrated targeted proteomics successfully, their possibilities in discovery-oriented proteomics have not yet been thoroughly explored. Our objective was to determine what are the extra costs and what optimization and adjustments to a conventional-flow LC-MS system must be undertaken to identify a comparable number of proteins as can be identified on a nanoLC-MS system. We demonstrate that the amount of a complex tryptic digest needed for comparable proteome coverage can be roughly 5-fold greater, providing the column dimensions are properly chosen, extra-column peak dispersion is minimized, column temperature and flow rate are set to levels appropriate for peptide separation, and the composition of mobile phases is fine-tuned. Indeed, we identified 2 835 proteins from 2 mu g of HeLa cells tryptic digest separated during a 60 min gradient at 68 mu L/min on a 1.0 mm x 250 mm column held at 55 degrees C and using an aqua-acetonitrile mobile phases containing 0.1% formic acid, 0.4% acetic acid, and 3% dimethyl sulfoxide. Our results document that conventional-flow LC-MS is an attractive alternative for bottom-up exploratory proteomics.
Czech name
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Czech description
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Classification
Type
J<sub>imp</sub> - Article in a specialist periodical, which is included in the Web of Science database
CEP classification
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OECD FORD branch
10406 - Analytical chemistry
Result continuities
Project
Result was created during the realization of more than one project. More information in the Projects tab.
Continuities
P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)<br>I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace
Others
Publication year
2018
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Name of the periodical
Analytical Chemistry
ISSN
0003-2700
e-ISSN
—
Volume of the periodical
90
Issue of the periodical within the volume
8
Country of publishing house
US - UNITED STATES
Number of pages
9
Pages from-to
5381-5389
UT code for WoS article
000430512200061
EID of the result in the Scopus database
2-s2.0-85045630358