Engineering a tunable micropattern‐array assay to sort single extracellular vesicles and particles to detect RNA and protein in situ

Author:

Zhang Jingjing1,Rima Xilal Y.1ORCID,Wang Xinyu1,Nguyen Luong T. H.1ORCID,Huntoon Kristin23,Ma Yifan1,Palacio Paola Loreto4,Nguyen Kim Truc1,Albert Karunya1,Duong‐Thi Minh‐Dao1,Walters Nicole1,Kwak Kwang Joo5,Yoon Min Jin1,Li Hong1,Doon‐Ralls Jacob1,Hisey Colin L.1,Lee Daeyong2,Wang Yifan6,Ha Jonghoon6,Scherler Kelsey7,Fallen Shannon7,Lee Inyoul7,Palmer Andre F.1,Jiang Wen6,Magaña Setty M.4,Wang Kai7,Kim Betty Y. S.23,Lee L. James15,Reátegui Eduardo18ORCID

Affiliation:

1. William G. Lowrie Department of Chemical and Biomolecular Engineering The Ohio State University Columbus Ohio USA

2. Department of Neurosurgery The University of Texas MD Anderson Cancer Center Houston Texas USA

3. The Brain Tumor Center The University of Texas MD Anderson Cancer Center Houston Texas USA

4. Department of Pediatrics, Division of Neurology Nationwide Children's Hospital Columbus Ohio USA

5. Spot Biosystems Ltd. Palo Alto California USA

6. Department of Radiation Oncology The University of Texas Southwestern Medical Center Dallas Texas USA

7. Institute for Systems Biology Seattle Washington USA

8. Comprehensive Cancer Center The Ohio State University Columbus Ohio USA

Abstract

AbstractThe molecular heterogeneity of extracellular vesicles (EVs) and the co‐isolation of physically similar particles, such as lipoproteins (LPs), confounds and limits the sensitivity of EV bulk biomarker characterization. Herein, we present a single‐EV and particle (siEVP) protein and RNA assay (siEVPPRA) to simultaneously detect mRNAs, miRNAs, and proteins in subpopulations of EVs and LPs. The siEVPPRA immobilizes and sorts particles via positive immunoselection onto micropatterns and focuses biomolecular signals in situ. By detecting EVPs at a single‐particle resolution, the siEVPPRA outperformed the sensitivities of bulk‐analysis benchmark assays for RNA and protein. To assess the specificity of RNA detection in complex biofluids, EVs from various glioma cell lines were processed with small RNA sequencing, whereby two mRNAs and two miRNAs associated with glioblastoma multiforme (GBM) were chosen for cross‐validation. Despite the presence of single‐EV‐LP co‐isolates in serum, the siEVPPRA detected GBM‐associated vesicular RNA profiles in GBM patient siEVPs. The siEVPPRA effectively examines intravesicular, intervesicular, and interparticle heterogeneity with diagnostic promise.

Funder

National Institutes of Health

National Center for Advancing Translational Sciences

Publisher

Wiley

Subject

Cell Biology,Histology

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