From shotgun to targeted proteomics: rapid Scout-MRM assay development for monitoring potential immunomarkers in Dreissena polymorpha

Autor: Alain Geffard, Christine Almunia, Jérôme Lemoine, Arnaud Salvador, Julien Faugere, Mélissa Palos-Ladeiro, Maxime Leprêtre, Jean Armengaud
Přispěvatelé: Stress Environnementaux et BIOsurveillance des milieux aquatiques (SEBIO), Institut National de l'Environnement Industriel et des Risques (INERIS)-Université de Reims Champagne-Ardenne (URCA)-Université Le Havre Normandie (ULH), Normandie Université (NU)-Normandie Université (NU)-SFR Condorcet, Université de Reims Champagne-Ardenne (URCA)-Centre National de la Recherche Scientifique (CNRS)-Université de Reims Champagne-Ardenne (URCA)-Centre National de la Recherche Scientifique (CNRS), ANABIO-MS - Analyse biomoléculaire par spectrométrie de masse - Biological Analysis by Mass Spectrometry, Institut des Sciences Analytiques (ISA), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), We thank the University of Reims Champagne-Ardenne for its financial support (project PROMETHEE)., Université de Reims Champagne-Ardenne (URCA)-Université de Picardie Jules Verne (UPJV)-Centre National de la Recherche Scientifique (CNRS)-Université de Reims Champagne-Ardenne (URCA)-Université de Picardie Jules Verne (UPJV)-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS)
Jazyk: angličtina
Rok vydání: 2020
Předmět:
Zdroj: Analytical and Bioanalytical Chemistry
Analytical and Bioanalytical Chemistry, 2020, 412 (26), pp.7333-7347. ⟨10.1007/s00216-020-02868-2⟩
Analytical and Bioanalytical Chemistry, Springer Verlag, 2020, 412 (26), pp.7333-7347. ⟨10.1007/s00216-020-02868-2⟩
ISSN: 1618-2642
1618-2650
DOI: 10.1007/s00216-020-02868-2⟩
Popis: This work benefitted from the French GDR "Aquatic Ecotoxicology" framework which is aimed at fostering stimulating scientific discussions and collaborations for more integrative approaches.; International audience; A highly multiplexed liquid chromatography mass spectrometry-multiple reaction monitoring (MRM)-based assay has been developed for evaluating 107 candidate immune biomarkers in both hemocytes and plasma of the zebra musselDreissena polymorpha. The Scout-MRM strategy was employed for the first time, shortening the implementation of a targeted MRM bottom-up proteomics assay using selected immune protein-related peptides identified by shotgun discovery proteogenomics. This strategy relies on spiking scout peptides during the discovery phase and using them to build and deploy the MRM targeted proteomics method. It proved to be highly relevant, since about 90% of the targeted peptides and proteins were monitored and rapidly measured in both hemocyte and plasma samples. The sample preparation protocol was optimized by evaluating the digestion efficiency of tryptic peptides over time. The accuracy and precision of 50 stable isotope-labeled peptides were evaluated for use as internal standards. Finally, the specificity of the transitions was thoroughly assessed to ensure the reliable measurement of protein biomarkers. Several analytical and biological validation criteria were evaluated across hemocytes and plasma samples exposed ex vivo to biological contaminants, resulting in the validation of two Scout-MRM assays for the relative quantitation of 85 and 89 proteins in hemocytes and plasma, respectively.
Databáze: OpenAIRE
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