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Comparing Approaches to Normalize, Quantify, and Characterize Urinary Extracellular Vesicles.

Charles J Blijdorp1, Omar A Z Tutakhel2,3, Thomas A Hartjes4

  • 1Division of Nephrology and Transplantation, Department of Internal Medicine, Erasmus Medical Center, University Medical Center Rotterdam, Rotterdam, The Netherlands.

Journal of the American Society of Nephrology : JASN
|March 30, 2021
PubMed
Summary
This summary is machine-generated.

Urinary extracellular vesicle (uEV) concentration correlates with creatinine, simplifying spot urine normalization. Dilute urine and Tamm-Horsfall protein affect particle detection methods.

Keywords:
aquaporin-2biomarkercreatinineexosomesnormalizationosmolalityparticlestetraspaninurinary extracellular vesiclesuromodulin

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Area of Science:

  • Biochemistry
  • Nanotechnology
  • Urology

Background:

  • Urinary extracellular vesicles (uEVs) show promise for biomarker discovery.
  • Standardized methods for uEV analysis in spot urines are lacking.
  • Challenges include normalization, quantification, and characterization.

Purpose of the Study:

  • To evaluate methods for uEV analysis in spot urines.
  • To investigate the correlation between uEVs and creatinine.
  • To understand factors affecting uEV quantification.

Main Methods:

  • Analysis of urine samples from healthy subjects and kidney disease patients.
  • Quantification using Nanoparticle Tracking Analysis (NTA), time-resolved fluorescence immunoassay (TR-FIA), and EVQuant.
  • Investigation of water loading effects and Tamm-Horsfall protein (THP) interference.

Main Results:

  • High correlation between uEV and creatinine concentrations in all study groups.
  • Water loading increased particle detection by NTA, linked to hypotonicity and reduced THP polymerization.
  • SDS improved uEV integrity and aquaporin-2 detection; reverse topology uEVs and nephron segment-specific EVs were observed.

Conclusions:

  • Urine creatinine concentration can potentially replace uEV quantification for normalization in spot urines.
  • Tamm-Horsfall protein interferes with NTA; dilute urine yields larger uEVs.
  • Detergents enhance intracellular epitope recognition, and specific markers capture nephron segment-specific EVs.