title: A modified precipitation method to isolate urinary exosomes.
abstract: Identification of biomarkers that allow early detection of kidney diseases in urine and plasma has been an area of active interest for several years. Urinary exosome vesicles, 40-100 nm in size, are released into the urine under normal conditions by cells from all nephron segments and may contain protein, mRNA and microRNA representative of their cell type of origin. Under conditions of renal dysfunction or injury, exosomes may contain altered proportions of these components, which may serve as biomarkers for disease. There are currently several methods available for isolation of urinary exosomes, and we have previously conducted an experimental comparison of each of these approaches, including three based on ultracentrifugation, one using a nanomembrane ultrafiltration concentrator, one using a commercial precipitation reagent and one using a modification of the precipitation technique using ExoQuick reagent that we developed in our laboratory. We found the modified precipitation method produced the highest yield of exosome particles, miRNA, and mRNA, making this approach suitable for the isolation of exosomes for subsequent RNA profiling. We conclude that the modified exosome precipitation method offers a quick, scalable, and effective alternative for the isolation of exosomes from urine. In this report, we describe our modified precipitation technique using ExoQuick reagent for isolating exosomes from human urine.
authors: Diabetes, Cardiovascular and Metabolic Diseases Division, Translational Genomics Research Institute (TGen).
authors: Rupesh
authors: R
authors: Kanchi Ravi
authors: Diabetes, Cardiovascular and Metabolic Diseases Division, Translational Genomics Research Institute (TGen).
authors: Mahdieh
authors: M
authors: Khosroheidari
authors: Diabetes, Cardiovascular and Metabolic Diseases Division, Translational Genomics Research Institute (TGen); jdistefano@tgen.org.
authors: Johanna K
authors: JK
authors: DiStefano
mesh_terms: D015415:Biomarkers / Q000652:urine; D055354:Exosomes / Q000737:chemistry*; D005590:Fractional Precipitation / Q000379:methods*; D006801:Humans; D035683:MicroRNAs / Q000302:isolation & purification / Q000652:urine*; D012333:RNA, Messenger / Q000302:isolation & purification / Q000652:urine*; D014461:Ultracentrifugation / Q000379:methods; D014462:Ultrafiltration; D016482:Urinalysis / Q000379:methods
chemical_list: D015415:Biomarkers; D035683:MicroRNAs; D012333:RNA, Messenger
references: Kidney Int. 2012 Jun;81(12):1263-72
references: 22418980
references: Proc Natl Acad Sci U S A. 2004 Sep 7;101(36):13368-73
references: 15326289
references: Curr Protoc Cell Biol. 2006 Apr;Chapter 3:Unit 3.22
references: 18228490
references: Kidney Int. 2010 Oct;78(8):810-6
references: 20686450
references: Methods Mol Biol. 2010;641:89-99
references: 20407943
references: Kidney Int. 2010 Jul;78(2):191-9
references: 20428099
references: Ren Fail. 2007;29(6):693-8
references: 17763164
references: J Am Soc Nephrol. 2009 Feb;20(2):363-79
references: 19056867
references: Am J Physiol Renal Physiol. 2007 May;292(5):F1657-61
references: 17229675
references: Kidney Int. 2006 Apr;69(8):1471-6
references: 16501490
references: Proteomics Clin Appl. 2012 Jun;6(5-6):268-78
references: 22641613
references: Kidney Int. 2012 Nov;82(9):1024-32
references: 22785172