Competitive magneto enzyme-linked miRNA assay for quantification of microRNAs in exosomes
Bernuz, Mireia (Universitat Autònoma de Barcelona. Institut de Biotecnologia i de Biomedicina "Vicent Villar Palasí")
Martí, Mercè (Universitat Autònoma de Barcelona. Departament de Biologia Cel·lular, de Fisiologia i d'Immunologia)
Pividori, María Isabel (Universitat Autònoma de Barcelona. Departament de Química)

Data:	2026
Resum:	MicroRNAs (miRNAs) are non-coding, short RNA sequences involved in gene regulation. Although intracellular molecules, miRNAs are also found in biological fluids, where they serve as promising diagnostic and prognostic biomarkers. In liquid biopsies, miRNAs are protected from RNase activity through association with proteins or as part of the exosomal cargo. Current methods for miRNA detection are technically demanding and resource intensive. Here, we present a competitive magneto enzyme-linked miRNA (ELmiRNA) assay for the direct quantification of miRNAs in exosomes. The assay is based on a DNA scaffold containing two miRNA templates, each hybridized to a complementary double-tagged probe and anchored via a poly(A) tail to oligo(dT)-magnetic particles. Upon incubation with miRNA-containing samples, displacement of the tagged probes occurs, enabling indirect quantification through enzymatic labeling. As proof of concept, miRNA-21 and miRNA-27a, both upregulated in breast cancer, were quantified in MCF-7 and MDA-MB-231 cells and their derived exosomes. The assay achieved limits of detection of 1. 5 nM (9. 6 pg μL) and 2. 7 nM (16. 2 pg μL) for miRNA-21 and miRNA-27a, respectively, and yielded 10-30 copies per exosome, in agreement with RT-qPCR validation. The ELmiRNA assay combines sensitivity and selectivity with high-throughput analysis using a routine laboratory filter-based microplate reader. This approach represents a simplified, amplification-free alternative to RT-qPCR for miRNA quantification in exosomes, suitable for routine implementation in standard clinical and research laboratories.
Ajuts:	Agencia Estatal de Investigación PID2022-136453OB-I00 Agencia Estatal de Investigación PDC2022-133363-I00 Generalitat de Catalunya 2021/SGR-00124 Generalitat de Catalunya 2021/SGR-00740
Nota:	Altres ajuts: acords transformatius de la UAB
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	miRNA ; Magneto enzyme-linked assay ; Exosome analysis ; Biomarkers ; Magnetic particles ; Liquid biopsy
Publicat a:	Talanta, Vol. 302 (May 2026) , art. 129431, ISSN 1873-3573

DOI: 10.1016/j.talanta.2026.129431

9 p, 3.6 MB

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