Dataset Information

Different EV enrichment methods suitable for clinical settings yield different subpopulations of urinary extracellular vesicles from human samples.

ABSTRACT: Urine sample analysis is irreplaceable as a non-invasive method for disease diagnosis and follow-up. However, in urine samples, non-degraded protein and RNA may be only found in urinary extracellular vesicles (uEVs). In recent years, various methods of uEV enrichment using low volumes of urine and unsophisticated equipment have been developed, with variable success. We compared the results of the differential ultracentrifugation procedure with 4 of these methods. The methods tested were a lectin-based purification, Exoquick (System Biosciences), Total Exosome Isolation from Invitrogen and an in-house modified procedure employing the Exosomal RNA Kit from Norgen Biotek Corp. The analysis of selected gene transcripts and protein markers of extracellular vesicles (EVs) revealed that each method isolates a different mixture of uEV protein markers. In our conditions, the extraction with Norgen's reagent achieved the best performance in terms of gene transcript and protein detection and reproducibility. By using this method, we were able to detect alterations of EVs protein markers in urine samples from prostate cancer adenoma patients. Taken together, our results show that the isolation of uEVs is feasible from small volumes of urine and avoiding ultracentrifugation, making easier the analysis in a clinical facility. However, caution should be taken in the selection of the enrichment method since they have a differential affinity for protein uEVs markers and by extension for different subpopulation of EVs.

SUBMITTER: Royo F

PROVIDER: S-EPMC4759834 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Different EV enrichment methods suitable for clinical settings yield different subpopulations of urinary extracellular vesicles from human samples.

Royo Felix F Zuñiga-Garcia Patricia P Sanchez-Mosquera Pilar P Egia Ainara A Perez Amparo A Loizaga Ana A Arceo Raquel R Lacasa Isabel I Rabade Ainara A Arrieta Edurne E Bilbao Roberto R Unda Miguel M Carracedo Arkaitz A Falcon-Perez Juan M JM

Journal of extracellular vesicles 20160215

Urine sample analysis is irreplaceable as a non-invasive method for disease diagnosis and follow-up. However, in urine samples, non-degraded protein and RNA may be only found in urinary extracellular vesicles (uEVs). In recent years, various methods of uEV enrichment using low volumes of urine and unsophisticated equipment have been developed, with variable success. We compared the results of the differential ultracentrifugation procedure with 4 of these methods. The methods tested were a lectin ...[more]

PMID: 26895490

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Project description:Extracellular vesicles (EVs) play a key role in cellular communication both in physiological conditions and in pathologies such as cancer. Emerging evidence has shown that EVs are active carriers of molecular cargo (e.g. protein and nucleic acids) and a powerful source of biomarkers and targets. While recent studies on EV-associated DNA (EV-DNA) in human biofluids have generated a large amount of data, there is currently no database that catalogues information on EV-DNA. To fill this gap, we have manually curated a database of EV-DNA data derived from human biofluids (liquid biopsy) and in-vitro studies, called the Extracellular Vesicle-Associated DNA Database (EV-ADD). This database contains validated experimental details and data extracted from peer-reviewed published literature. It can be easily queried to search for EV isolation methods and characterization, EV-DNA isolation techniques, quality validation, DNA fragment size, volume of starting material, gene names and disease context. Currently, our database contains samples representing 23 diseases, with 13 different types of EV isolation techniques applied on eight different human biofluids (e.g. blood, saliva). In addition, EV-ADD encompasses EV-DNA data both representing the whole genome and specifically including oncogenes, such as KRAS, EGFR, BRAF, MYC, and mitochondrial DNA (mtDNA). An EV-ADD data metric system was also integrated to assign a compliancy score to the MISEV guidelines based on experimental parameters reported in each study. While currently available databases document the presence of proteins, lipids, RNA and metabolites in EVs (e.g. Vesiclepedia, ExoCarta, ExoBCD, EVpedia, and EV-TRACK), to the best of our knowledge, EV-ADD is the first of its kind to compile all available EV-DNA datasets derived from human biofluid samples. We believe that this database provides an important reference resource on EV-DNA-based liquid biopsy research, serving as a learning tool and to showcase the latest developments in the EV-DNA field. EV-ADD will be updated yearly as newly published EV-DNA data becomes available and it is freely available at www.evdnadatabase.com.

Dataset Information

Different EV enrichment methods suitable for clinical settings yield different subpopulations of urinary extracellular vesicles from human samples.

Publications

Different EV enrichment methods suitable for clinical settings yield different subpopulations of urinary extracellular vesicles from human samples.

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