Dataset Information

Improving peptide fragment ion detection using travelling wave ion mobility spectrometry with signal enhancement MSE (seMSE)

ABSTRACT: Recently, a solution to inherent poor ion beam sampling of time-of-flight (TOF) mass analyzers was proposed by restricting detection to ions expected at specific arrival times after being separated in a travelling wave ion mobility spectrometry (TWIMS) cell. Here, we show that this approach can be applied to data independent acquisition (DIA) using a so-called signal enhancement MSE (seMSE) experiment, which was compared to two different TWIMS-DDA and three different TWIMS-MSE methods with respect to the number of peptide identifications, the spectral quality of supporting peptide spec-tra matches, and, more importantly, fragment ion sensitivity. Comparison of fragment ion summed areas showed that seM-SE showed 6.8 to 11.5-fold improvement in fragment ion detection when compared to other MSE methods and although co-fragmentation of co-eluting peptides limited the seMSE peptide identifications it was possible to consistently detect peptides of interest by integrating DDA and MSE results within Skyline.

ORGANISM(S): Oryctolagus Cuniculus Escherichia Coli Bos Taurus Saccharomyces Cerevisiae

SUBMITTER: Juan Camilo Rojas Echeverri

PROVIDER: PXD030431 | panorama | Mon Aug 01 00:00:00 BST 2022

REPOSITORIES: PanoramaPublic

ACCESS DATA

Publications

Evaluating Peptide Fragment Ion Detection Using Traveling Wave Ion Mobility Spectrometry with Signal-Enhanced MSE (SEMSE).

Rojas Echeverri Juan Camilo JC Volke Daniela D Milkovska-Stamenova Sanja S Hoffmann Ralf R

Analytical chemistry 20220729 31

The inherent poor sampling of fragment ions in time-of-flight mass analyzers was recently improved for data-dependent acquisition (DDA) by considering their drift times in traveling wave ion mobility spectrometry (TWIMS). Here, we extend this TWIMS-DDA approach to the data-independent acquisition (DIA) mode MSE to improve the signal intensities of fragment ions by providing improved ion beam sampling efficiency, which we termed therefore signal-enhanced MSE (SEMSE</sup ...[more]

PMID: 35904512

Similar Datasets

Project description:Although hepatocellular carcinoma (HCC) has been subjected to continuous investigation and its symptoms are well-known, early stage diagnosis of this disease remains difficult and the survival rate after diagnosis is typically very low (3−5%). Early and accurate detection of metabolic changes in the sera of patients with liver cirrhosis can help improve the prognosis of HCC and lead to a better understanding of its mechanism at the molecular level, thus providing patients with in-time treatment of the disease. In this study, we compared metabolite levels in sera of 40 HCC patients and 49 cirrhosis patients from Egypt by using ultraperformance liquid chromatography coupled with quadrupole time-of-flight mass spectrometer (UPLC-QTOF MS). Following data preprocessing, the most relevant ions in distinguishing HCC cases from cirrhotic controls are selected by statistical methods. Putative metabolite identifications for these ions are obtained through mass-based database search. The identities of some of the putative identifications are verified by comparing their MS/MS fragmentation patterns and retention times with those from authentic compounds. Finally, the serum samples are reanalyzed for quantitation of selected metabolites as candidate biomarkers of HCC. This quantitation was performed using isotope dilution by selected reaction monitoring (SRM) on a triple quadrupole linear ion trap (QqQLIT) coupled to UPLC. Statistical analysis of the UPLC-QTOF data identified 274 monoisotopic ion masses with statistically significant differences in ion intensities between HCC cases and cirrhotic controls. Putative identifications were obtained for 158 ions by mass based search against databases. We verified the identities of selected putative identifications including glycholic acid (GCA), glycodeoxycholic acid (GDCA), 3β, 6β-dihydroxy-5β-cholan-24-oic acid, oleoyl carnitine, and Phe-Phe. SRM-based quantitation confirmed significant differences between HCC and cirrhotic controls in metabolite levels of bile acid metabolites, long chain carnitines and small peptide. Our study provides useful insight into appropriate experimental design and computational methods for serum biomarker discovery using LC−MS/MS based metabolomics. This study has led to the identification of candidate biomarkers with significant changes in metabolite levels between HCC cases and cirrhotic controls. This is the first MS-based metabolic biomarker discovery study on Egyptian subjects that led to the identification of candidate metabolites that discriminate early stage HCC from patients with liver cirrhosis.

Project description:The sensitivity of single-cell proteomics (SCP) has increased dramatically in recent years due to advances in experimental design, sample preparation, separations and mass spectrometry instrumentation. However, further increasing the sensitivity of SCP methods and instrumentation will enable the study of proteins within single cells that are expressed at copy numbers too small to be measured by current methods. Here we further increase SCP sensitivity by combining efficient nanoPOTS sample preparation and ultra-low-flow liquid chromatography with a newly developed data acquisition and analysis scheme termed wide window acquisition (WWA) to extend the achievable proteome coverage for single cells to >3,000 for single-cell data analyses. WWA is based on data-dependent acquisition (DDA) but employs larger precursor isolation windows to intentionally co-isolate and co-fragment additional precursors along with the selected precursor. The resulting chimeric MS2 spectra are then resolved using the CHIMERYS search engine within Proteome Discoverer 3.0. Compared to standard DDA workflows, WWA employing isolation windows of 8-12 Th increases peptide and proteome coverage by ~28% and ~39%, respectively. For a 40 min LC gradient operated at ~15 nL/min, we identified an average of 2,150 proteins per single-cell-sized aliquots of protein digest directly from MS2 spectra, which increased to an average 3,524 proteins including proteins identified with MS1-level feature matching. Reducing the active gradient to 20 min resulted in only a 10% decrease in proteome coverage. We also compared performance of WWA with DIA underperformed relative to WWA in terms of proteome coverage, especially with faster separations. Average proteome coverage for single HeLa and K562 cells was respectively 1,758 and 1,642 based on MS2 identifications with 1% false discovery rate and 3042 and 2891 with MS1 feature matching. As such, WWA combined with efficient sample preparation and rapid separations extends the depths of the proteome that can be studied at the single-cell level.

Dataset Information

Improving peptide fragment ion detection using travelling wave ion mobility spectrometry with signal enhancement MSE (seMSE)

Publications

Evaluating Peptide Fragment Ion Detection Using Traveling Wave Ion Mobility Spectrometry with Signal-Enhanced MS<sup>E</sup> (SEMS<sup>E</sup>).

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets