Comparison of Extracellular Vesicle Isolation Methods for miRNA Sequencing
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.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2023 |
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| Erschienen: |
2023 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:24 |
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| Enthalten in: |
International journal of molecular sciences - 24(2023), 15 vom: 29. Juli |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Llorens-Revull, Meritxell [VerfasserIn] |
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| Links: |
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| Themen: |
Diagnostic biomarker |
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| Anmerkungen: |
Date Completed 14.08.2023 Date Revised 14.08.2023 published: Electronic Citation Status MEDLINE |
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| doi: |
10.3390/ijms241512183 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM360690742 |
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| 245 | 1 | 0 | |a Comparison of Extracellular Vesicle Isolation Methods for miRNA Sequencing |
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| 500 | |a Citation Status MEDLINE | ||
| 520 | |a 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 | ||
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| 700 | 1 | |a Martínez-González, Brenda |e verfasserin |4 aut | |
| 700 | 1 | |a Quer, Josep |e verfasserin |4 aut | |
| 700 | 1 | |a Esteban, Juan Ignacio |e verfasserin |4 aut | |
| 700 | 1 | |a Núñez-Moreno, Gonzalo |e verfasserin |4 aut | |
| 700 | 1 | |a Mínguez, Pablo |e verfasserin |4 aut | |
| 700 | 1 | |a Burgui, Idoia |e verfasserin |4 aut | |
| 700 | 1 | |a Ramos-Ruíz, Ricardo |e verfasserin |4 aut | |
| 700 | 1 | |a Soria, María Eugenia |e verfasserin |4 aut | |
| 700 | 1 | |a Rico, Angie |e verfasserin |4 aut | |
| 700 | 1 | |a Riveiro-Barciela, Mar |e verfasserin |4 aut | |
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| 700 | 1 | |a Piron, María |e verfasserin |4 aut | |
| 700 | 1 | |a Corrales, Irene |e verfasserin |4 aut | |
| 700 | 1 | |a Borràs, Francesc E |e verfasserin |4 aut | |
| 700 | 1 | |a Rodríguez-Frías, Francisco |e verfasserin |4 aut | |
| 700 | 1 | |a Rando, Ariadna |e verfasserin |4 aut | |
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| 700 | 1 | |a Costafreda, Maria Isabel |e verfasserin |4 aut | |
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