Extracellular Vesicle Capture by AnTibody of CHoice and Enzymatic Release (EV-CATCHER): A customizable purification assay designed for small-RNA biomarker identification and evaluation of circulating small-EVs

J Extracell Vesicles. 2021 Jun;10(8):e12110. doi: 10.1002/jev2.12110. Epub 2021 Jun 3.


Circulating nucleic acids, encapsulated within small extracellular vesicles (EVs), provide a remote cellular snapshot of biomarkers derived from diseased tissues, however selective isolation is critical. Current laboratory-based purification techniques rely on the physical properties of small-EVs rather than their inherited cellular fingerprints. We established a highly-selective purification assay, termed EV-CATCHER, initially designed for high-throughput analysis of low-abundance small-RNA cargos by next-generation sequencing. We demonstrated its selectivity by specifically isolating and sequencing small-RNAs from mouse small-EVs spiked into human plasma. Western blotting, nanoparticle tracking, and transmission electron microscopy were used to validate and quantify the capture and release of intact small-EVs. As proof-of-principle for sensitive detection of circulating miRNAs, we compared small-RNA sequencing data from a subset of small-EVs serum-purified with EV-CATCHER to data from whole serum, using samples from a small cohort of recently hospitalized Covid-19 patients. We identified and validated, only in small-EVs, hsa-miR-146a and hsa-miR-126-3p to be significantly downregulated with disease severity. Separately, using convalescent sera from recovered Covid-19 patients with high anti-spike IgG titers, we confirmed the neutralizing properties, against SARS-CoV-2 in vitro, of a subset of small-EVs serum-purified by EV-CATCHER, as initially observed with ultracentrifuged small-EVs. Altogether our data highlight the sensitivity and versatility of EV-CATCHER.

Keywords: TEM; exosome purification; extracellular vesicles; micro‐RNA profiling; sequencing.

MeSH terms

  • Animals
  • Bodily Secretions / chemistry
  • COVID-19 / blood
  • COVID-19 / physiopathology
  • Chlorocebus aethiops
  • Circulating MicroRNA
  • Extracellular Vesicles / chemistry*
  • High-Throughput Nucleotide Sequencing
  • Humans
  • Immunologic Techniques / methods*
  • MCF-7 Cells
  • Mice
  • RAW 264.7 Cells
  • Severity of Illness Index
  • Vero Cells


  • Circulating MicroRNA