Dataset Information

Extending the Separation Space with Trapped Ion Mobility Spectrometry Improves the Accuracy of Isobaric Tag-based Quantitation in Proteomic LC/MS/MS

ABSTRACT: Two-dimensional separation by nano-LC and trapped ion mobility spectrometry (TIMS) prior to Q-TOF tandem mass spectrometry significantly improves the accuracy of isobaric tag-based quantitation in proteome analysis without the need for additional measurement time for TIMS insertion. The obtained peak capacity of up to 3,300 per hour in LC/TIMS reduced the co-isolation of precursor ions at the quadrupole analyzer, resulting in more accurate ratios of reporter ions derived from isobaric tags in product ion spectra obtained at the TOF analyzer. We also found that TIMS with a narrow quadrupole isolation window could reduce the ratio compression effect at least as effectively as the synchronous precursor selection method using MS3 scans without compromising sensitivity or coverage. Our results suggest that the short-gradient LC/TIMS/Q/TOF system is an excellent platform for high-throughput proteomics studies.

ORGANISM(S): Cellular Organisms

SUBMITTER: Yasushi Ishihama

PROVIDER: PXD018650 | JPOST Repository | Fri May 22 00:00:00 BST 2020

REPOSITORIES: jPOST

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	190209ko04HeLa_EColi_TMT_TIMS_ColRF500_IW2_Slot1-01_1_1725.tdf	Other
	190209ko04HeLa_EColi_TMT_TIMS_ColRF500_IW2_Slot1-01_1_1725.tdf_bin	Other
	190209ko04HeLa_EColi_TMT_TIMS_ColRF500_IW2_Slot1-01_1_1725_5.1.201.dat	Other
	190209ko04HeLa_EColi_TMT_TIMS_ColRF500_IW2_Slot1-01_1_1725_5.1.201.mgf	Mgf
	190209ko05HeLa_EColi_TMT_TIMS_ColRF500_IW1_Slot1-01_1_1726.tdf	Other

Items per page:

1 - 5 of 17

Publications

Extending the Separation Space with Trapped Ion Mobility Spectrometry Improves the Accuracy of Isobaric Tag-Based Quantitation in Proteomic LC/MS/MS.

Ogata Kosuke K Ishihama Yasushi Y

Analytical chemistry 20200528 12

Two-dimensional separation by nano-LC and trapped ion mobility spectrometry (TIMS) prior to Q/TOF tandem mass spectrometry significantly improves the accuracy of isobaric tag-based quantitation in proteome analysis without the need for additional measurement time for TIMS insertion between LC and Q/TOF MS. The obtained peak capacity of up to 3300 h<sup>-1</sup> in LC/TIMS reduced the coisolation of precursor ions at the quadrupole analyzer, resulting in more accurate ratios of reporter ions deri ...[more]

PMID: 32441512

Similar Datasets

Project description:Data independent acquisition-mass spectrometry (DIA-MS) coupled with liquid chromatography is a promising approach for rapid, automatic sampling of MS/MS data in untargeted metabolomics. However, wide isolation windows in DIA-MS generate MS/MS spectra containing a mixed population of fragment ions together with their precursor ions. This precursor-fragment ion map in a comprehensive MS/MS spectral library is crucial for relative quantification of fragment ions uniquely representative of each precursor ion. However, existing reference libraries are not sufficient for this purpose since the fragmentation patterns of small molecules can vary in different instrument setups. Here we developed a bioinformatics workflow called MetaboDIA to build customized MS/MS spectral libraries using a user's own data dependent acquisition (DDA) data and to perform MS/MS-based quantification with DIA data, thus complementing conventional MS1-based quantification. MetaboDIA also allows users to build a spectral library directly from DIA data in studies of a large sample size. Using a marine algae data set, we show that quantification of fragment ions extracted with a customized MS/MS library can provide as reliable quantitative data as the direct quantification of precursor ions based on MS1 data. To test its applicability in complex samples, we applied MetaboDIA to a clinical serum metabolomics data set, where we built a DDA-based spectral library containing consensus spectra for 1829 compounds. We performed fragment ion quantification using DIA data using this library, yielding sensitive differential expression analysis. </br></br> Serum metabolome of 40 age-related macular degeneration patients and 20 control samples was analyzed using untargeted mass spectrometry. We used data dependent acquisition data to build a MS/MS spectral assay library for more than 1,000 compounds and performed targeted extraction of MS2 ion chromatograms from data independent acquisition analysis.