Isolation and profiling of microRNA-containing exosomes from human bile

Ling Li, Klaus B. Piontek, Vivek Kumbhari, Masaharu Ishida, Florin M. Selaru

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Exosome research in the last three years has greatly extended the scope towards identification and characterization of biomarkers and their therapeutic uses. Exosomes have recently been shown to contain microRNAs (miRs). MiRs themselves have arisen as valuable biomarkers for diagnostic purposes. As specimen collection in clinics and hospitals is quite variable, miRNA isolation from whole bile varies substantially. To achieve robust, accurate and reproducible miRNA profiles from collected bile samples in a simple manner required the development of a highquality protocol to isolate and characterize exosomes from bile. The method requires several centrifugations and a filtration step with a final ultracentrifugation step to pellet the isolated exosomes. Electron microscopy, Western blots, flow cytometry and multi-parameter nanoparticle optical analysis, where available, are crucial characterization steps to validate the quality of the exosomes. For the isolation of miRNA from these exosomes, spiking the lysate with a non-specific, synthetic miRNA from a species like Caenorhabditis elegans, i.e., Cel-miR-39, is important for normalization of RNA extraction efficiency. The isolation of exosome from bile fluid following this method allows the successful miRNA profiling from bile samples stored for several years at-80 °C.

Original languageEnglish
Article numbere54036
JournalJournal of Visualized Experiments
Volume2016
Issue number112
DOIs
Publication statusPublished - 2016 Jun 13

Keywords

  • Bile
  • Electron microscopy
  • Exosomes
  • Extracellular vesicles
  • Issue 112
  • Medicine
  • MicroRNA
  • Molecular biology
  • Nanoparticle optical analysis
  • Western Blot

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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