MassIVE MSV000093468

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Towards an ultimate solution for peptide retention time prediction: the effect of column temperature on separation selectivity

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Description

The effect of the column temperature on the selectivity of reversed-phase peptide separation in standard separation settings applied in bottom-up proteomics has been evaluated. Tryptic digests of A549, MCF7, Jurkat and HCT116 human cells in non-labeled and TMT-labeled formats have been analyzed by 1D LC-MS/MS at four different column temperature settings: 25, 35, 45 and 55 C. To increase the size of retention datasets available for retention time prediction modelling, 2D LC-MS/MS acquisitions were also performed on a Jurkat digest (both non-labeled and TMT-modified). The first dimension separation featured high pH RPLC separation (XTerra, 1x100 mm column, pH 10, 2 % per minute acetonitrile gradient). Fourteen (non-labeled) and eighteen (TMT) concatenated fractions were analyzed using RPLC-MS settings identical to that of the 1D experiments. The second dimension of RP chromatographic separation employed the Thermo Fisher Scientific EASY Spray column (PepMap, RSLC C18, 2mm, 100-angstrom, 75mm x 50cm), operated at four temperatures with a gradient of 1% to 50% B (A: 100% water, B: 80:20 ACN: water, both containing 0.1% formic acid) in 107 min, followed by a 1 min rapid increase to 100%B and 12 minute wash. The total acquisition time for each sample/fraction was 120 min. [doi:10.25345/C50R9MF37] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: peptide separation ; temperature impact ; predictive modeling ; machine learning ; chromatography fundmanetal studies

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Principal Investigators:
(in alphabetical order) 		Oleg Krokhin, University of Manitoba, Canada
Submitting User: vspicer1
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