Skip to main content
bioRxiv
  • Home
  • About
  • Submit
  • ALERTS / RSS
Advanced Search
New Results

An optimized method for the isolation of urinary extracellular vesicles for molecular phenotyping: detection of biomarkers for radiation exposure

View ORCID ProfileCharles P. Hinzman, Meth Jayatilake, Sunil Bansal, Brian L. Fish, Yaoxiang Li, Yubo Zhang, Shivani Bansal, Michael Girgis, Anton Iliuk, Xiao Xu, Jose A. Fernandez, John H. Griffin, Elizabeth A Ballew, Keith Unger, Marjan Boerma, Meetha Medhora, Amrita K. Cheema
doi: https://doi.org/10.1101/2022.01.28.477909
Charles P. Hinzman
1Department of Biochemistry and Molecular & Cellular Biology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Charles P. Hinzman
Meth Jayatilake
2Department of Oncology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Sunil Bansal
2Department of Oncology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Brian L. Fish
3Department of Radiation Oncology, Medical College of Wisconsin, Milwaukee, WI 53226
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yaoxiang Li
2Department of Oncology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yubo Zhang
2Department of Oncology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Shivani Bansal
2Department of Oncology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Michael Girgis
2Department of Oncology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Anton Iliuk
4Tymora Analytical Operations, West Lafayette, IN 47906
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Xiao Xu
5Department of Molecular Medicine, Scripps Research Institute, La Jolla, CA 92037
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jose A. Fernandez
5Department of Molecular Medicine, Scripps Research Institute, La Jolla, CA 92037
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
John H. Griffin
5Department of Molecular Medicine, Scripps Research Institute, La Jolla, CA 92037
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Elizabeth A Ballew
6Department of Radiation Medicine, MedStar Georgetown University Hospital, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Keith Unger
6Department of Radiation Medicine, MedStar Georgetown University Hospital, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Marjan Boerma
7University of Arkansas for Medical Sciences, Department of Pharmaceutical Sciences, Division of Radiation Health, Little Rock, AK 72205
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Meetha Medhora
3Department of Radiation Oncology, Medical College of Wisconsin, Milwaukee, WI 53226
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Amrita K. Cheema
1Department of Biochemistry and Molecular & Cellular Biology, Georgetown University Medical Center, Washington, DC 20007
2Department of Oncology, Georgetown University Medical Center, Washington, DC 20007
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • For correspondence: akc27@georgetown.edu
  • Abstract
  • Full Text
  • Info/History
  • Metrics
  • Supplementary material
  • Preview PDF
Loading

ABSTRACT

Background Urinary extracellular vesicles (EVs) are a source of biomarkers with broad potential applications across clinical research, including monitoring radiation exposure. A key limitation to their implementation is minimal standardization in EV isolation and analytical analysis methods. Further, most urinary EV isolation protocols necessitate large volumes of sample. This study aimed to compare and optimize isolation and analytical methods for EVs from small volumes of urine.

Methods 3 EV isolation methods were compared: ultracentrifugation, magnetic bead-based, and size-exclusion chromatography from 0.5 mL or 1 mL of rat and human urine. EV yield and mass spectrometry signals (Q-ToF and Triple Quad) were evaluated from each method. Metabolomic profiling was performed on EVs isolated from the urine of rats exposed to ionizing radiation 1-, 14-, 30- or 90-days post-exposure, and human urine from patients receiving thoracic radiotherapy for the treatment of lung cancer pre- and post-treatment.

Results Size-exclusion chromatography is the preferred method for EV isolation from 0.5 mL of urine. Mass spectrometry-based metabolomic analyses of EV cargo identified biochemical changes induced by radiation, including altered nucleotide, folate, and lipid metabolism. We have provided standard operating procedures for implementation of these methods in other laboratories.

Conclusions We demonstrate that EVs can be isolated from small volumes of urine and analytically investigated for their biochemical contents to detect radiation induced metabolomic changes. These findings lay a groundwork to develop future methods to monitor response to radiotherapy and can be extended to an array of molecular phenotyping studies aimed at characterizing EV cargo.

Competing Interest Statement

The authors have declared no competing interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-ND 4.0 International license.
Back to top
PreviousNext
Posted January 28, 2022.
Download PDF

Supplementary Material

Email

Thank you for your interest in spreading the word about bioRxiv.

NOTE: Your email address is requested solely to identify you as the sender of this article.

Enter multiple addresses on separate lines or separate them with commas.
An optimized method for the isolation of urinary extracellular vesicles for molecular phenotyping: detection of biomarkers for radiation exposure
(Your Name) has forwarded a page to you from bioRxiv
(Your Name) thought you would like to see this page from the bioRxiv website.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
An optimized method for the isolation of urinary extracellular vesicles for molecular phenotyping: detection of biomarkers for radiation exposure
Charles P. Hinzman, Meth Jayatilake, Sunil Bansal, Brian L. Fish, Yaoxiang Li, Yubo Zhang, Shivani Bansal, Michael Girgis, Anton Iliuk, Xiao Xu, Jose A. Fernandez, John H. Griffin, Elizabeth A Ballew, Keith Unger, Marjan Boerma, Meetha Medhora, Amrita K. Cheema
bioRxiv 2022.01.28.477909; doi: https://doi.org/10.1101/2022.01.28.477909
Digg logo Reddit logo Twitter logo Facebook logo Google logo LinkedIn logo Mendeley logo
Citation Tools
An optimized method for the isolation of urinary extracellular vesicles for molecular phenotyping: detection of biomarkers for radiation exposure
Charles P. Hinzman, Meth Jayatilake, Sunil Bansal, Brian L. Fish, Yaoxiang Li, Yubo Zhang, Shivani Bansal, Michael Girgis, Anton Iliuk, Xiao Xu, Jose A. Fernandez, John H. Griffin, Elizabeth A Ballew, Keith Unger, Marjan Boerma, Meetha Medhora, Amrita K. Cheema
bioRxiv 2022.01.28.477909; doi: https://doi.org/10.1101/2022.01.28.477909

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Subject Area

  • Cancer Biology
Subject Areas
All Articles
  • Animal Behavior and Cognition (3609)
  • Biochemistry (7590)
  • Bioengineering (5533)
  • Bioinformatics (20833)
  • Biophysics (10347)
  • Cancer Biology (7998)
  • Cell Biology (11663)
  • Clinical Trials (138)
  • Developmental Biology (6619)
  • Ecology (10227)
  • Epidemiology (2065)
  • Evolutionary Biology (13648)
  • Genetics (9557)
  • Genomics (12860)
  • Immunology (7932)
  • Microbiology (19576)
  • Molecular Biology (7678)
  • Neuroscience (42193)
  • Paleontology (309)
  • Pathology (1259)
  • Pharmacology and Toxicology (2208)
  • Physiology (3272)
  • Plant Biology (7064)
  • Scientific Communication and Education (1295)
  • Synthetic Biology (1953)
  • Systems Biology (5435)
  • Zoology (1119)