Data independent acquisition (DIA) strategies have emerged as a powerful alternative to classic data dependent acquisition (DDA) in shotgun proteomics. DIA offers systematic measurement of all peptide ions regardless of their intensity by co-fragmenting all co-eluting peptides in broader precursor isolation windows. This provides a wider dynamic range of the proteomes analyzed, improved reproducibility for identification and enables better sensitivity and accuracy for quantification.

While modern mass spectrometers have developed towards faster scanning speeds and enhanced selectivity by ion mobility, it is indeed the elaborate processing algorithms for identification, that has made the most significant contributions to the field over the last couple of years. The analysis of DIA data has historically been difficult and driven by specialists in the field. One of the most important requirements to perform a DIA analysis has been to generate a project specific library, which is made from extensive offline fractionation with the same LC gradient and column as used in the actual DIA experiment.

It is now also possible to search DIA data without the need for DDA based spectral libraries, which is a game-changer, facilitating simpler and more powerful workflows. Further implementation of deep-learning based models has significantly contributed to increased proteome coverage. These major developments over the last years have positioned DIA as an important acquisition strategy and the preferred method in many labs.

The trend continues

This is a especially positive, as DIA is the ideal match for short and standardized gradients on the Evosep One. It has been demonstrated that more than 5000 proteins can be identified in just 5 min using dia-PASEF with the fast 200 samples per day method. This proteome coverage is increased to nearly 7000 proteins when extending the gradient time using the 60 samples per day method. With such a coverage obtained in short time, it is possible to gain relevant biological insight for discovery proteomics with large cohorts of clinical samples.

Webinar: Unleash the power of DIA

The steep development over the last years, have positioned DIA as an important acquisition strategy to generate record high identifications with short gradients on the Evosep One. Join this webinar to explore how our customers have used Evosep One to generate record high IDs per minute for large-scale sample cohorts.

You can also read more about the research published using DIA methodologies from our users here.

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MORE RESEARCH ON DIA

Here you can see publications available using DIA featuring Evosep One. For a full overview of publications published using the Evosep One Technology visit our Literature room here

TitleSubjectMaterialYearSummaryEvosep methodMS instrumentationLearn More
Data independent acquisition enables deep and fast label-free dynamic organellar mapping, 2021

This publication by the Borner group, describes a DIA workflow to establish organellar maps, which combines cell fractionation and shotgun proteomics into a profiling analysis of subcellular localization. They apply this to HeLa cells and investigate changes in response to starvation/disruption of lysosomal pH.

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Unleash the power of DIA2022

In this webinar we dive into the exciting research possibilities that comes by using DIA combined with short gradients with Evosep One.

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Ultra-high sensitivity mass spectrometry quantifies single-cell proteome changes upon perturbation, , 2020

In this publication, the Mann group in Münich describes their innovative workflow for single-cell proteomics. They apply a nearly loss-less miniaturized sample preparation workflow to our robust Whisper 40SPD method and combine it with diaPASEF on a modified timsTOF mass spectrometer for a total of 420 FACS sorted single cells.

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DebaryOmics: an integrative –omics study to understand the halophilic behaviour of Debaryomyces hansenii, , , 2021

This publication by the Martinez group at the Technical University of Denmark, presents the first global phosphoproteomics analysis in Debaryomyces hansenii, a non-conventional yeast considered to be a well-suited option for a number of different industrial bioprocesses.

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Targeted Workflows with Evosep One, , 2021

During this webinar, Michael MacCoss (University of Washington) and Vivian Delcourt (GIE-LCH) share how they use the Evosep One for targeted workflows.

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Plasma Proteomics with the Evosep One, , 2021

In this webinar you will learn how the Evosep One, with high reproducibility and robustness, is ideally suited for analysis of large sample cohorts of blood plasma.

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AlphaTims: Indexing trapped ion mobility spectrometry – time of flight data for fast and easy accession and visualization, , 2021

This publication by the Mann group, introduces the open-source AlphaTims package with a stand-alone graphical user interface or as part of their AlphaPept ecosystem. With this package, unprocessed data are indexed, accessed, and visualized by a combination of tools.

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Proteomics of resistance to Notch1 inhibition in acute lymphoblastic leukemia reveals targetable kinase signatures, , , , , 2021

In this publication, the Olsen group highlights the potential of proteomics to dissect alterations in cellular signaling and identify druggable pathways in cancer. They identify protein kinase C delta (PKCδ) inhibition as a strategy to overcome resistance to Notch1 inhibition in T-cell acute lymphoblastic leukemia.

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Unleashing the power of DIA combined with short gradients using Evosep One, 2021

This webinar is part of our Evosep webinar series and hosted by Nicolai Bache, Evosep. Florian Meier and Vadim Demichev discuss their use of the Evosep One, using novel DIA methodologies.

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Unleashing the true power of DIA/SWATH data acquisition with short gradients – Rapid profiling of pre-clinical models and proteome dynamics, , 2019

Achieving sufficient statistical power for the analysis of a comprehensive set of target proteins requires processing of a large number of samples. With finite resources, researchers face a difficult choice: should they aim at deeper proteomic coverage or higher sample throughput?

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Standardized workflow for precise mid- and high-throughput proteomics of blood biofluids, , , 2021

In this seminar from the Canadian National Proteomics Network (CNPN) meeting 2021, Angela McArdle presents a standardized workflow for precise high-throughput proteomics of blood biofluids. They established optimal conditions for sample preparation and DIA analysis in plasma, then automated and adapted this for depleted plasma and whole blood.

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Integrative analysis of cell state changes in lung fibrosis with peripheral protein biomarkers, , , 2021

In this study, the Theis and Schiller groups at the Helmholtz Zentrum München, investigated the correspondence of cell state changes in diseased organs to peripheral protein signatures in pulmonary fibrosis patient cohorts. From plasma proteome profiling of more than 122 patients, they propose CRTAC1 protein levels in plasma as a novel biomarker.

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Covid-19 research with Evosep One, , 2021

This webinar is part of our Evosep webinar series and hosted by Nicolai Bache, Evosep. Angela McArdle and Mathias Trost discuss their use of the Evosep One technology in their development of rapid Covid-19 assays.

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SPIN – Species by Proteome Investigation, 2021

This publication from the Olsen Group, introduces “Species by Proteome INvestigation” (SPIN), a proteomics workflow capable of querying over 150 mammalian species 200 times a day with the Evosep One. Genetic species determination is an indispensable tool in forensics, archaeology, ecology, and food authentication.

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Spatial-proteomics reveal in-vivo phospho-signaling dynamics at subcellular resolution, , , 2021

This publication from the Olsen group at the Novo Nordisk Foundation Center for Protein Research, describes a high-throughput workflow based on sequential cell fractionation to profile the global proteome and phospho-proteome dynamics across six distinct subcellular fractions.

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High throughput proteome and phosphorpoteome sample processing coupled to fast gradient DIA, , , , , , , 2020

High throughput workflows for proteomics

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New Orbitrap Exploris 480 Mass Spectrometer Coupled with Evosep One, , , 2020

Brochure of Exploris 480 with Evosep data from piece no 1

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A Novel LC System Embeds Analytes in Pre-formed Gradients for Rapid, Ultra-Robust Proteomics, , , 2018

Description of Evosep

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High dynamic range proteome analysis with BoxCar DIA and super-resolution Orbitrap mass spectrometry, , 2020

BoxCar DIA

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Increasing proteome coverage using cysteine-specific DIA Mass spectrometry – Cys-DIA, 2020

DIA on cysteine-specific peptides

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Improving proteome coverage and peptide identification rates in short LC gradients, , , , , 2020, ,

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High-throughput proteomics with Evosep One, , , , , 2020, ,

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Rapid proteome analyses using the Evosep One, , , 2018

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Using Artificial Intelligence on Ultrafast LC-MSMS-DIA runs for Bacterial Identification in Urine, , 2019

View the recording of our HUPO 2019 lunch seminar, where Florence Roux-Dalvai from Québec Research Center, Canada presents a new strategy for bacterial species identification in urinary tract infection using artificial intelligence on ultrafast LCMS DIA runs.  

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High-throughput 4D-Proteomics – Application of dia-PASEF® and the Evosep One for short gradients, 2020

diaPASEF app note  

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Oncogenic Mutations Rewire Signaling Pathways by Switching Protein Recruitment to Phosphotyrosine Sites, , , , 2019

In this publication, the Olsen Group have combined DIA with short LC gradients to describe how cancer mutations close to tyrosine phosphorylation sites in the EGF receptor rewire signaling pathways by switching protein interactors. The performed 1200 pulldown experiments in just 20 days of LC-MS instrument time.

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Multi-level proteomics reveals host-perturbation strategies of SARS-CoV-2 and SARS-CoV, , , 2020

This publication describes the molecular functions of viral proteins and their interactions with the host proteome of SARS-CoV-2. The impact of viral infection on the proteome and phosphoproteome were analyzed in a time-resolved manner by DIA. The analysis revealed key pathways perturbed during the infection identifying potential vulnerable points of SARS-CoV-2.

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A paired liver biopsy and plasma proteomics study reveals circulating biomarkers for alcohol-related liver disease, , , 2020

This publication from Matthias Mann groups in Copenhagen and Münich describes a paired liver biopsy and plasma proteomics study, which make use of Boxcar DIA scanning for a large clinical cohort of nearly 600 patients. They have developed a machine learning model based on their biomarker panel, which for the first time outperforms existing tests, …

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Increasing Throughput: From Pre-Clinical Models To Protein Complexes, 2019

Comparison study with >400 samples, same coverage in shorter time

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A Compact Quadrupole-Orbitrap Mass Spectrometer With Faims Interface Improves Proteome Coverage In Short Lc Gradients, , , , , , , , , 2020

Test of Orbitrap Exploris with DIA and TMT, proteome and phospho.

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