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Comparison of three methods for isolation of urinary microvesicles to identify biomarkers of nephrotic syndrome.

Abstract：

:Urinary microvesicles, such as 40-100 nm exosomes and 100-1000 nm microparticles, contain many proteins that may serve as biomarkers of renal disease. Microvesicles have been isolated by ultracentrifugation or nanomembrane ultrafiltration from normal urine; however, little is known about the efficiency of these methods in isolating microvesicles from patients with nephrotic-range proteinuria. Here we compared three techniques to isolate microvesicles from nephrotic urine: nanomembrane ultrafiltration, ultracentrifugation, and ultracentrifugation followed by size-exclusion chromatography (UC-SEC). Highly abundant urinary proteins were still present in sufficient quantity after ultrafiltration or ultracentrifugation to blunt detection of less abundant microvesicular proteins by MALDI-TOF-TOF mass spectrometry. The microvesicular markers neprilysin, aquaporin-2, and podocalyxin were highly enriched following UC-SEC compared with preparations by ultrafiltration or ultracentrifugation alone. Electron microscopy of the UC-SEC fractions found microvesicles of varying size, compatible with the presence of both exosomes and microparticles. Thus, UC-SEC following ultracentrifugation to further enrich and purify microparticles facilitates the search for prognostic biomarkers that might be used to predict the clinical course of nephrotic syndrome.

journal_name

Kidney Int

journal_title

Kidney international

authors

Rood IM,Deegens JK,Merchant ML,Tamboer WP,Wilkey DW,Wetzels JF,Klein JB

doi

10.1038/ki.2010.262

subject

Has Abstract

pub_date

2010-10-01 00:00:00

pages

810-6

issue

8

eissn

0085-2538

issn

1523-1755

pii

S0085-2538(15)54636-X

journal_volume

78

pub_type

杂志文章

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