title: A simplified method to recover urinary vesicles for clinical applications, and sample banking.
abstract: Urinary extracellular vesicles provide a novel source for valuable biomarkers for kidney and urogenital diseases: Current isolation protocols include laborious, sequential centrifugation steps which hampers their widespread research and clinical use. Furthermore, large individual urine sample volumes or sizable target cohorts are to be processed (e.g. for biobanking), the storage capacity is an additional problem. Thus, alternative methods are necessary to overcome such limitations. We have developed a practical vesicle isolation technique to yield easily manageable sample volumes in an exceptionally cost efficient way to facilitate their full utilization in less privileged environments and maximize the benefit of biobanking. Urinary vesicles were isolated by hydrostatic dialysis with minimal interference of soluble proteins or vesicle loss. Large volumes of urine were concentrated up to 1/100 of original volume and the dialysis step allowed equalization of urine physico-chemical characteristics. Vesicle fractions were found suitable to any applications, including RNA analysis. In the yield, our hydrostatic filtration dialysis system outperforms the conventional ultracentrifugation-based methods and the labour intensive and potentially hazardous step of ultracentrifugations are eliminated. Likewise, the need for trained laboratory personnel and heavy initial investment is avoided. Thus, our method qualifies as a method for laboratories working with urinary vesicles and biobanking.
authors.affiliation: Centre for BioAnalytical Sciences (CBAS), Dublin City University, Republic of Ireland.
authors.forename: Luca
authors.initials: L
authors.lastname: Musante
authors.affiliation: Centre for BioAnalytical Sciences (CBAS), Dublin City University, Republic of Ireland.
authors.forename: Dorota
authors.initials: D
authors.lastname: Tataruch
authors.affiliation: Centre for BioAnalytical Sciences (CBAS), Dublin City University, Republic of Ireland.
authors.forename: Dongfeng
authors.initials: D
authors.lastname: Gu
authors.affiliation: Centre for BioAnalytical Sciences (CBAS), Dublin City University, Republic of Ireland.
authors.forename: Alberto
authors.initials: A
authors.lastname: Benito-Martin
authors.affiliation: Centre for BioAnalytical Sciences (CBAS), Dublin City University, Republic of Ireland.
authors.forename: Giulio
authors.initials: G
authors.lastname: Calzaferri
authors.affiliation: National Institute for Cellular Biotechnology, Dublin City University, Republic of Ireland.
authors.forename: Sinead
authors.initials: S
authors.lastname: Aherne
authors.affiliation: Centre for BioAnalytical Sciences (CBAS), Dublin City University, Republic of Ireland.
authors.forename: Harry
authors.initials: H
authors.lastname: Holthofer
mesh_terms: D000328:Adult; D018070:Biological Specimen Banks; D055252:Cell-Derived Microparticles; D005260:Female; D005831:Genital Diseases, Female / Q000652:urine*; D005832:Genital Diseases, Male / Q000652:urine*; D006801:Humans; D008297:Male; D011309:Preservation, Biological / Q000379:methods*; D014570:Urologic Diseases / Q000652:urine*