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Adv."],"published-print":{"date-parts":[[2026,8,21]]},"abstract":"<jats:p>\n                    Extracellular vesicle\u2013associated microRNAs (EV-miRNAs) hold promise for liquid biopsy, but their clinical utility is hindered by EV heterogeneity. Here, we report DEEPER, a DNA-encoded orthogonal recognition platform for selective barcoding of tumor-derived EV subpopulations and in situ miRNA profiling. DEEPER uses four aptamers targeting Cluster of Differentiation 44 (CD44), Epidermal Growth Factor Receptor\u200c (EGFR), Human Epidermal Growth Factor Receptor 2\u200c (HER2), and \u200cProgrammed Death-Ligand 1(PD-L1) to generate orthogonal barcodes that distinguish 15 EV subpopulations. Barcoded EVs then undergo targeted membrane fusion with liposome probes, enabling sensitive in situ analysis of subpopulation-specific miRNAs. In a 60-sample cohort of patients with gastric cancer (GC) and healthy donors (HDs), DEEPER identified a high-performing signature, GC-EV miRNA Prime, comprising EGFR\n                    <jats:sup>+<\/jats:sup>\n                    , EGFR\n                    <jats:sup>+<\/jats:sup>\n                    HER2\n                    <jats:sup>+<\/jats:sup>\n                    , and EGFR\n                    <jats:sup>+<\/jats:sup>\n                    HER2\n                    <jats:sup>+<\/jats:sup>\n                    CD44\n                    <jats:sup>+<\/jats:sup>\n                    EVs, which achieved 98.3% diagnostic accuracy. The same profiling framework also assessed tumor invasiveness with 95.5% accuracy. Mechanistic studies further showed that these EV-miRNAs contribute to tumor progression. In summary, DEEPER not only confirmed the clinical application value of EV-miRNA\u2013based liquid biopsy but also provided unique insights through the EV subpopulations into the mechanisms driving cancer development.\n                  <\/jats:p>","DOI":"10.1126\/sciadv.aef8133","type":"journal-article","created":{"date-parts":[[2026,8,21]],"date-time":"2026-08-21T17:58:41Z","timestamp":1787335121000},"update-policy":"https:\/\/doi.org\/10.34133\/aaas_crossmark","source":"Crossref","is-referenced-by-count":1,"title":["Orthogonal DNA barcoding enables subpopulation-resolved extracellular vesicle miRNA profiling"],"prefix":"10.1126","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0009-0007-5610-9186","authenticated-orcid":true,"given":"Ye","family":"Zhang","sequence":"first","affiliation":[{"name":"Department of Laboratory Medicine, Guangdong Provincial Key Laboratory of Precision Medical Diagnostics, Guangdong Engineering and Technology Research Center for Rapid Diagnostic Biosensors, Guangdong Provincial Key Laboratory of Single-cell and Extracellular Vesicles, Nanfang Hospital, Southern Medical University, Guangzhou 510515, P. R. China."},{"name":"Guangdong Provincial Clinical Research Center for Laboratory Medicine, Guangzhou, China."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0009-1275-0240","authenticated-orcid":true,"given":"Yitong","family":"Zhu","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Guangdong Provincial Key Laboratory of Precision Medical Diagnostics, Guangdong Engineering and Technology Research Center for Rapid Diagnostic Biosensors, Guangdong Provincial Key Laboratory of Single-cell and Extracellular Vesicles, Nanfang Hospital, Southern Medical University, Guangzhou 510515, P. R. China."},{"name":"Guangdong Provincial Clinical Research Center for Laboratory Medicine, Guangzhou, China."},{"name":"Medical Research Center of Nanfang Hospital, Nanfang Hospital, Guangdong Provincial Key Laboratory of Proteomics, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, P. R. China."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-1513-3757","authenticated-orcid":true,"given":"Yu","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Guangdong Provincial Key Laboratory of Precision Medical Diagnostics, Guangdong Engineering and Technology Research Center for Rapid Diagnostic Biosensors, Guangdong Provincial Key Laboratory of Single-cell and Extracellular Vesicles, Nanfang Hospital, Southern Medical University, Guangzhou 510515, P. R. China."},{"name":"Guangdong Provincial Clinical Research Center for Laboratory Medicine, Guangzhou, China."},{"name":"Medical Research Center of Nanfang Hospital, Nanfang Hospital, Guangdong Provincial Key Laboratory of Proteomics, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, P. R. China."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0005-6346-1001","authenticated-orcid":true,"given":"Shijin","family":"Peng","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Guangdong Provincial Key Laboratory of Precision Medical Diagnostics, Guangdong Engineering and Technology Research Center for Rapid Diagnostic Biosensors, Guangdong Provincial Key Laboratory of Single-cell and Extracellular Vesicles, Nanfang Hospital, Southern Medical University, Guangzhou 510515, P. R. China."},{"name":"Guangdong Provincial Clinical Research Center for Laboratory Medicine, Guangzhou, China."},{"name":"Medical Research Center of Nanfang Hospital, Nanfang Hospital, Guangdong Provincial Key Laboratory of Proteomics, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, P. R. China."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Diheng","family":"Wu","sequence":"additional","affiliation":[{"name":"School of Biomedical Engineering, Southern Medical University, Guangzhou, Guangdong 510515, P. R. China."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenbin","family":"Li","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Guangdong Provincial Key Laboratory of Precision Medical Diagnostics, Guangdong Engineering and Technology Research Center for Rapid Diagnostic Biosensors, Guangdong Provincial Key Laboratory of Single-cell and Extracellular Vesicles, Nanfang Hospital, Southern Medical University, Guangzhou 510515, P. R. China."},{"name":"Guangdong Provincial Clinical Research Center for Laboratory Medicine, Guangzhou, China."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tingting","family":"Ji","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, Guangdong Provincial Key Laboratory of Precision Medical Diagnostics, Guangdong Engineering and Technology Research Center for Rapid Diagnostic Biosensors, Guangdong Provincial Key Laboratory of Single-cell and Extracellular Vesicles, Nanfang Hospital, Southern Medical University, Guangzhou 510515, P. R. 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