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We characterized the surface epitopes of extracellular vesicles (EVs) from plasma HIV\/HCV-coinfected patients and their smRNA cargo profile, by comparing different isolation procedures. Six EVs isolation procedures were compared: ultracentrifugation, and five different polyethylene glycol-based methods (commercial, combined with a column purification step and two custom); and two RNA commercial kits (phenol and non-phenol based) were used. High-throughput sequencing of smRNAs was performed. Exosomal surface epitopes were analyzed by the MACSPlex Exosome Kit. Four miRNAs displayed differences among protocols (hsa-miR-205-5p and hsa-let-7a\/b\/f-5p). The selection of RNA isolation kit impacted on the detection of miRNAs and other smRNAs, where the phenol-based RNA isolation kit performed acceptably. EVs surface was enriched with HLA-DR\/DP\/DQ, CD81, and CD8. There were three liver-specific miRNAs\u00a0overexpressed (let-7a-5p, miR-21-5p and hsa-miR-122-5p), thus, EVs cargo might reflect liver disease evolution. Other smRNAs such as piwi-interacting RNAs were also detected for the first time. Custom polyethylene glycol precipitation-based methods combined with an RNA phenol-based kit yielded the higher number of smRNAs for EVs isolated from plasma HIV\/HCV patients.<\/jats:p>","DOI":"10.1038\/s41598-020-67935-1","type":"journal-article","created":{"date-parts":[[2020,7,7]],"date-time":"2020-07-07T10:04:08Z","timestamp":1594116248000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":32,"title":["Comparison of methods and characterization of small RNAs from plasma extracellular vesicles of HIV\/HCV coinfected patients"],"prefix":"10.1038","volume":"10","author":[{"given":"Elena","family":"Mart\u00ednez-Gonz\u00e1lez","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"\u00d3scar","family":"Brochado-Kith","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alicia","family":"G\u00f3mez-Sanz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Luz","family":"Mart\u00edn-Carbonero","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ma \u00c1ngeles","family":"Jimenez-Sousa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paula","family":"Mart\u00ednez-Rom\u00e1n","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Salvador","family":"Resino","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ver\u00f3nica","family":"Briz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amanda","family":"Fern\u00e1ndez-Rodr\u00edguez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2020,7,7]]},"reference":[{"key":"67935_CR1","unstructured":"1Organization, W. 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