Comparison of Extracellular Vesicle Isolation Methods for miRNA Sequencing.

Autor:	Llorens-Revull M; Liver Diseases-Viral Hepatitis, Liver Unit, Vall d'Hebron Institut de Recerca (VHIR), Vall d'Hebron Barcelona Hospital Campus, Passeig Vall d'Hebron 119-129, 08035 Barcelona, Spain.; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Biochemistry and Molecular Biology Department, Universitat Autònoma de Barcelona (UAB), Campus de la UAB, Plaza Cívica, 08193 Bellaterra, Spain., Martínez-González B; Department of Clinical Microbiology, IIS-Fundación Jiménez Díaz, UAM, Av. Reyes Católicos 2, 28040 Madrid, Spain.; Department of Molecular and Cell Biology, Centro Nacional de Biotecnología (CNB-CSIC), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, 28049 Madrid, Spain., Quer J; Liver Diseases-Viral Hepatitis, Liver Unit, Vall d'Hebron Institut de Recerca (VHIR), Vall d'Hebron Barcelona Hospital Campus, Passeig Vall d'Hebron 119-129, 08035 Barcelona, Spain.; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Biochemistry and Molecular Biology Department, Universitat Autònoma de Barcelona (UAB), Campus de la UAB, Plaza Cívica, 08193 Bellaterra, Spain., Esteban JI; Liver Diseases-Viral Hepatitis, Liver Unit, Vall d'Hebron Institut de Recerca (VHIR), Vall d'Hebron Barcelona Hospital Campus, Passeig Vall d'Hebron 119-129, 08035 Barcelona, Spain.; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Biochemistry and Molecular Biology Department, Universitat Autònoma de Barcelona (UAB), Campus de la UAB, Plaza Cívica, 08193 Bellaterra, Spain., Núñez-Moreno G; Department of Genetics & Genomics, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain.; CIBER de Enfermedades Raras (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Bioinformatics Unit, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain., Mínguez P; Department of Genetics & Genomics, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain.; CIBER de Enfermedades Raras (CIBERER), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Bioinformatics Unit, Instituto de Investigación Sanitaria-Fundación Jiménez Díaz University Hospital, Universidad Autónoma de Madrid (IIS-FJD, UAM), 28040 Madrid, Spain., Burgui I; Unidad de Genómica, 'Scientific Park of Madrid', Campus de Cantoblanco, 28049 Madrid, Spain., Ramos-Ruíz R; Unidad de Genómica, 'Scientific Park of Madrid', Campus de Cantoblanco, 28049 Madrid, Spain., Soria ME; Department of Clinical Microbiology, IIS-Fundación Jiménez Díaz, UAM, Av. Reyes Católicos 2, 28040 Madrid, Spain.; Centro de Biología Molecular 'Severo Ochoa' (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), 28049 Madrid, Spain., Rico A; Transfusion Safety Laboratory, Blood and Tissue Bank of Catalonia (BST), 08005 Barcelona, Spain., Riveiro-Barciela M; Liver Diseases-Viral Hepatitis, Liver Unit, Vall d'Hebron Institut de Recerca (VHIR), Vall d'Hebron Barcelona Hospital Campus, Passeig Vall d'Hebron 119-129, 08035 Barcelona, Spain.; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Biochemistry and Molecular Biology Department, Universitat Autònoma de Barcelona (UAB), Campus de la UAB, Plaza Cívica, 08193 Bellaterra, Spain., Sauleda S; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Transfusion Safety Laboratory, Blood and Tissue Bank of Catalonia (BST), 08005 Barcelona, Spain.; Transfusional Medicine, Vall d'Hebron Institut de Recerca (VHIR), Universitat Autònoma de Barcelona (UAB), 08035 Barcelona, Spain., Piron M; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Transfusion Safety Laboratory, Blood and Tissue Bank of Catalonia (BST), 08005 Barcelona, Spain.; Transfusional Medicine, Vall d'Hebron Institut de Recerca (VHIR), Universitat Autònoma de Barcelona (UAB), 08035 Barcelona, Spain., Corrales I; Transfusional Medicine, Vall d'Hebron Institut de Recerca (VHIR), Universitat Autònoma de Barcelona (UAB), 08035 Barcelona, Spain.; Congenital Coagulopathies Laboratory, Banc de Sang i Teixits (BST), 08005 Barcelona, Spain.; CIBER de Enfermedades Cardiovasculares (CIBERCV), Instituto de Salud Carlos III, 28029 Madrid, Spain., Borràs FE; REMAR-IVECAT Group, Germans Trias i Pujol Research Institute (IGTP), Can Ruti Campus, 08916 Badalona, Spain.; Nephrology Unit, Germans Trias i Pujol University Hospital, 08916 Badalona, Spain.; Department of Cell Biology, Physiology & Immunology, Universitat Autònoma de Barcelona (UAB), 08193 Barcelona, Spain., Rodríguez-Frías F; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Department of Basic Sciences, Universitat Internacional de Catalunya, Sant Cugat del Vallès, 08195 Barcelona, Spain.; Clinical Biochemistry Research Group, Vall d'Hebron Research Institute (VHIR), Biochemical Core Facilities, Vall d'Hebron University Hospital, Autonomous University Barcelona, 08035 Barcelona, Spain., Rando A; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Clinical Biochemistry Research Group, Vall d'Hebron Research Institute (VHIR), Biochemical Core Facilities, Vall d'Hebron University Hospital, Autonomous University Barcelona, 08035 Barcelona, Spain.; Microbiology Department Vall d'Hebron University Hospital, Barcelona Passeig Vall d'Hebron 119-129, 08035 Barcelona, Spain., Ramírez-Serra C; Clinical Biochemistry Research Group, Vall d'Hebron Research Institute (VHIR), Biochemical Core Facilities, Vall d'Hebron University Hospital, Autonomous University Barcelona, 08035 Barcelona, Spain., Camós S; Clinical Biochemistry Laboratory, ICS-IAS Girona Clinical Laboratory, Doctor Josep Trueta University Hospital, 17007 Girona, Spain., Domingo E; Centro de Biología Molecular 'Severo Ochoa' (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), 28049 Madrid, Spain., Bes M; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Transfusion Safety Laboratory, Blood and Tissue Bank of Catalonia (BST), 08005 Barcelona, Spain.; Transfusional Medicine, Vall d'Hebron Institut de Recerca (VHIR), Universitat Autònoma de Barcelona (UAB), 08035 Barcelona, Spain., Perales C; Department of Clinical Microbiology, IIS-Fundación Jiménez Díaz, UAM, Av. Reyes Católicos 2, 28040 Madrid, Spain.; Department of Molecular and Cell Biology, Centro Nacional de Biotecnología (CNB-CSIC), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, 28049 Madrid, Spain., Costafreda MI; Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, 28029 Madrid, Spain.; Enteric Virus Laboratory, Department of Genetics, Microbiology and Statistics, School of Biology, Institute of Nutrition and Safety, University of Barcelona (UB), 08007 Barcelona, Spain.
Jazyk:	angličtina
Zdroj:	International journal of molecular sciences [Int J Mol Sci] 2023 Jul 29; Vol. 24 (15). Date of Electronic Publication: 2023 Jul 29.
DOI:	10.3390/ijms241512183
Abstrakt:	MicroRNAs (miRNAs) encapsulated in extracellular vesicles (EVs) are potential diagnostic and prognostic biomarkers. However, discrepancies in miRNA patterns and their validation are still frequent due to differences in sample origin, EV isolation, and miRNA sequencing methods. The aim of the present study is to find a reliable EV isolation method for miRNA sequencing, adequate for clinical application. To this aim, two comparative studies were performed in parallel with the same human plasma sample: (i) isolation and characterization of EVs obtained using three procedures: size exclusion chromatography (SEC), iodixanol gradient (GRAD), and its combination (SEC+GRAD) and (ii) evaluation of the yield of miRNA sequences obtained using NextSeq 500 (Illumina) and three miRNA library preparation protocols: NEBNext, NEXTFlex, and SMARTer smRNA-seq. The conclusion of comparison (i) is that recovery of the largest amount of EVs and reproducibility were attained with SEC, but GRAD and SEC+GRAD yielded purer EV preparations. The conclusion of (ii) is that the NEBNext library showed the highest reproducibility in the number of miRNAs recovered and the highest diversity of miRNAs. These results render the combination of GRAD EV isolation and NEBNext library preparation for miRNA retrieval as adequate for clinical applications using plasma samples.
Databáze:	MEDLINE
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