Dataset Information

A Parallel Accumulation-Mobility Aligned Fragmentation Strategy Utilizing High-Resolution Ion Mobility for High Performance Proteomics Analysis: HYE Quantification Data Sets for PAMAF Mode

ABSTRACT: Here we present a parallel accumulation-mobility aligned fragmentation (PAMAF) mode that expands the capabilities of the MOBILion Systems MOBIE® high-resolution ion mobility (HRIM) system for mass spectrometry-based proteomics. The MOBIE system leverages structures for lossless ion manipulation (SLIM) to achieve HRIM separations within a compact footprint. Along with the HRIM separations, which increase the number of features identified per cycle, PAMAF enables alignment of fragment ions with their precursor ions without quadrupole isolation prior to fragmentation, avoiding the ion losses up to 99%. With PAMAF mode, ions can be accumulated in parallel to the ion mobility (IM) separation, achieving close to 100% ion utilization efficiency. Here, we demonstrate deep proteome coverage and robust LFQ accuracy and precision using PAMAF on a HRIM-QTOF platform.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human) Escherichia Coli Saccharomyces Cerevisiae (baker's Yeast)

TISSUE(S): Cell Suspension Culture, Cell Culture

SUBMITTER: Leonard Rorrer

LAB HEAD: Daniel DeBord

PROVIDER: PXD078350 | Pride | 2026-07-11

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	1000ng_HYE_MIX-A_htrms_files.zip	Other
	1000ng_HYE_MIX-A_mbi_files.zip	Other
	1000ng_HYE_MIX-B_htrms_files.zip	Other
	1000ng_HYE_MIX-B_mbi_files.zip	Other
	20260209_102620_1000ng_HYE_XIM_0.5_htrms_directDIA_Report_HYE_in_MSstats.zip	Other

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A Parallel Accumulation-Mobility Aligned Fragmentation Strategy Utilizing High-Resolution Ion Mobility for High-Performance Proteomics Analysis.

Rorrer Leonard C LC Deng Liulin L Royer Lauren L Uribe Isabel I Orsburn Benjamin C BC Bernhardt Oliver O Gandhi Tejas T Reiter Lukas L DeBord Daniel D

Molecular & cellular proteomics : MCP 20260622 7

Here we present a novel data-independent acquisition (DIA) mass spectrometry (MS) operating mode termed parallel accumulation-mobility aligned fragmentation (PAMAF) that offers enhanced speed and sensitivity of ion fragmentation analysis for discovery workflows such as bottom-up proteomics. This mode of operation leverages high-resolution ion mobility (HRIM) separation capabilities of the structures for lossless ion manipulation technology to achieve HRIM-based precursor isolation in place of tr ...[more]

PMID: 42331252

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Project description:Here we present a novel data independent acquisition mass spectrometry operating mode termed parallel accumulation-mobility aligned fragmentation (PAMAF) that offers enhanced speed and sensitivity for ion fragmentation analysis for nontargeted discovery workflows such as bottom-up proteomics. This mode of operation leverages high-resolution ion mobility (HRIM) separation capabilities of the structures for lossless ion manipulation (SLIM) technology to achieve HRIM-based precursor isolation in place of traditional quadrupole filtering approaches. This PAMAF mode of operation increases the number of features that can be identified per MS1/MS2 acquisition cycle, by employing mobility-based time alignment to associate fragment ions with their corresponding precursor ions. By using a high-speed, lossless separation technique for precursor isolation instead of the comparatively slow and wasteful quadrupole filtering method, we can avoid ion losses up to 99% while simultaneously increasing the rate at which precursor ions are sequentially fragmented and detected. Additionally, by storing ions in a trapping region while the previous packet of ions is being analyzed, the PAMAF mode achieves close to ~100% ion utilization efficiency. Benchmarking results of LC-PAMAF-MS analysis of a standard whole cell protein digest showed approximately 6x more protein group identifications compared to a standard data-dependent acquisition (DDA) analysis without HRIM on the same QTOF instrument. Quantitative evaluations demonstrated that PAMAF mode was capable of quantifying low abundance peptides, including those undetectable by DDA. Additionally, since precursor isolation in PAMAF mode is size-based rather than m/z-based, many coeluting isobars and isomers can be resolved prior to fragmentation to eliminate chimeric spectra that compromise identification accuracy. In this work we also explored the benefits of combining HRIM and quadrupole isolation to achieve maximum specificity. This approach, known as DIA-PAMAF mode, further reduces the frequency of chimeric fragmentation spectra, and enabled the detection of over 8,000 protein groups from a HeLa digest analysis. PAMAF mode brings a powerful new technique to the field of proteomics that has the potential to improve the sensitivity and selectivity of mass spectrometry-based proteomics.

Project description:We addressed the integrated analysis of mRNA and miRNA expression levels of Tg6799 AD model mice at 4 month and 8 months of age. Total 8 gene cluster modules for co-expression network were predicted from transcriptome data and 6 modules were show relation with AD or aging. We constructed early stage AD network using data integration between mRNA and miRNA profiles and predicted miRNAs strongly involved in module regulation. We found that ARRDC3 showed AD mutation dependent changes of expression and was related metabolic dysfunction in early stage AD. These results demonstrate that candidate genes on the simultaneous profiling of mRNA and miRNA expressions in genome wide can be used for the understanding of non-coding RNA related gene expression in early stage AD. We suggested that our results could be future candidate to be developed as early biomarkers in progressive AD pathology. This result can be used for the further application in neurodegenerative diseases. Tg6799 transgenic mice were purchased from The Jackson Laboratory (USA) and were housed under a 12h light-dark cycle with free access to food and water. Female Tg6799 mice are maintained until 4 months and 8 months of age (for littermate control: LM and mutant subjects: MT). RNA samples were isolated from hippocampus of mice using TRI-Reagent (Sigma-Aldrich, St. Louis, MO) according to the manufacturer’s instructions. Gene expression was analyzed with GeneChip® Mouse Genome 430 2.0 Arrays (Affymetrix, Santa Clara, CA), which is comprised of over 45,000 probe sets representing approximately 28,700 well-characterized mouse genes. The Ion Total RNA-Seq Kit v2 (Lifetechnologies, USA) was used for the preparation of micro RNA libraries according to the manufacturer's instructions. Total numbers of subject used are as followed: 1) LM 4 months : MT 4 months : LM 8 months : MT 8 months (2:4:2:4) for screening mRMA and miRNA, 2) LM 4 months : MT 4 months : LM 8 months : MT 8 months (4:4:4:4) for expression verification.

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