ArticleMolecular & cellular proteomics : MCP2016
Immobilized Metal Affinity Chromatography Coupled to Multiple Reaction Monitoring Enables Reproducible Quantification of Phospho-signaling.
Article in Molecular & cellular proteomics : MCP, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 26 papers.
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Who cites it
26 citing papers in PubMed, 49 citations in OpenAlex.
- RNA and phosphoprotein profiles of TP53- and PTEN-knockouts in MCF10A at baseline and responding to DNA damage.Scientific data · 2024Article
- Targeted Quantification of Protein Phosphorylation and Its Contributions towards Mathematical Modeling of Signaling Pathways.Molecules (Basel, Switzerland) · 2023Review
- Quantitative Cell Proteomic Atlas: Pathway-Scale Targeted Mass Spectrometry for High-Resolution Functional Profiling of Cell Signaling.Journal of proteome research · 2022Article
- Preserving the Phosphoproteome of Clinical Biopsies Using a Quick-Freeze Collection Device.Biopreservation and biobanking · 2022Article
- A highly multiplexed quantitative phosphosite assay for biology and preclinical studies.Molecular systems biology · 2021Article
- Targeted Mass Spectrometry Enables Quantification of Novel Pharmacodynamic Biomarkers of ATM Kinase Inhibition.Cancers · 2021Article
- Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling.Cell reports methods · 2021Article
- Loss of TGFβ signaling increases alternative end-joining DNA repair that sensitizes to genotoxic therapies across cancer types.Science translational medicine · 2021Article
- Targeted Mass Spectrometry Enables Multiplexed Quantification of Immunomodulatory Proteins in Clinical Biospecimens.Frontiers in immunology · 2021Article
- Deciphering MET-dependent modulation of global cellular responses to DNA damage by quantitative phosphoproteomics.Molecular oncology · 2020Article
- Clinical potential of mass spectrometry-based proteogenomics.Nature reviews. Clinical oncology · 2019Review
- Phosphopeptide Fragmentation and Site Localization by Mass Spectrometry: An Update.Analytical chemistry · 2019Review
- Quantitative targeted proteomic analysis of potential markers of tyrosine kinase inhibitor (TKI) sensitivity in EGFR mutated lung adenocarcinoma.Journal of proteomics · 2018Article
- Application of targeted mass spectrometry in bottom-up proteomics for systems biology research.Journal of proteomics · 2018Review
- Identification of Novel Physiological Substrates ofMolecular & cellular proteomics : MCP · 2018Article
- Therapy-induced stress response is associated with downregulation of pre-mRNA splicing in cancer cells.Genome medicine · 2018Article
- Targeted mass spectrometry enables robust quantification of FANCD2 mono-ubiquitination in response to DNA damage.DNA repair · 2018Article
- A Multiplexed Mass Spectrometry-Based Assay for Robust Quantification of Phosphosignaling in Response to DNA Damage.Radiation research · 2018Article
- Nonlinear Regression Improves Accuracy of Characterization of Multiplexed Mass Spectrometric Assays.Molecular & cellular proteomics : MCP · 2018Article
- Targeted Quantification of Phosphorylation Dynamics in the Context of EGFR-MAPK Pathway.Analytical chemistry · 2018Article
Corrections and comments
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Authors and funding
12 authors at 2 institutions in 1 country.
Funding
Abstract
A major goal in cell signaling research is the quantification of phosphorylation pharmacodynamics following perturbations. Traditional methods of studying cellular phospho-signaling measure one analyte at a time with poor standardization, rendering them inadequate for interrogating network biology and contributing to the irreproducibility of preclinical research. In this study, we test the feasibility of circumventing these issues by coupling immobilized metal affinity chromatography (IMAC)-based enrichment of phosphopeptides with targeted, multiple reaction monitoring (MRM) mass spectrometry to achieve precise, specific, standardized, multiplex quantification of phospho-signaling responses. A multiplex immobilized metal affinity chromatography- multiple reaction monitoring assay targeting phospho-analytes responsive to DNA damage was configured, analytically characterized, and deployed to generate phospho-pharmacodynamic curves from primary and immortalized human cells experiencing genotoxic stress. The multiplexed assays demonstrated linear ranges of ≥3 orders of magnitude, median lower limit of quantification of 0.64 fmol on column, median intra-assay variability of 9.3%, median inter-assay variability of 12.7%, and median total CV of 16.0%. The multiplex immobilized metal affinity chromatography- multiple reaction monitoring assay enabled robust quantification of 107 DNA damage-responsive phosphosites from human cells following DNA damage. The assays have been made publicly available as a resource to the community. The approach is generally applicable, enabling wide interrogation of signaling networks.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.