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You searched for: EV130016 (EV-TRACK ID)

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Experiment number
  • If needed, multiple experiments were identified in a single publication based on differing sample types, isolation protocols and/or vesicle types of interest.
Species
  • Species of origin of the EVs.
Isolation protocol
  • Gives a short, non-chronological overview of the different steps of the isolation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
Details EV-TRACK ID Experiment nr. Species Sample type Isolation protocol First author Year EV-METRIC
EV130016 1/1 Homo sapiens Urine dUC
Iodixanol-DG (valid.)
Raimondo F 2013 44%

Study summary

Full title
All authors
Raimondo F, Morosi L, Corbetta S, Chinello C, Brambilla P, Della Mina P, Villa A, Albo G, Battaglia C, Bosari S, Magni F, Pitto M
Journal
Mol Biosyst
Abstract
Renal cell carcinoma (RCC) accounts for about 3% of all human malignancies and its incidence is incr (show more...)Renal cell carcinoma (RCC) accounts for about 3% of all human malignancies and its incidence is increasing. There are no standard biomarkers currently used in the clinical management of patients with renal cell carcinoma. A promising strategy for new biomarker detection is comparative proteomics of urinary exosomes (UE), nanovesicles released by every epithelial cell facing the urinary space, enriched in renal proteins and excluding high-abundance plasmatic proteins, such as albumin. Aim of the work is to establish the protein profile of exosomes isolated from urines of RCC patient compared with control subjects. We enrolled 29 clear cell RCC patients and 23 control healthy subjects (CTRL), age and sex-matched, for urine collection and vesicle isolation by differential centrifugation. Such vesicles were morphologically and biochemically characterized and proved to share exosome properties. Proteomic analysis, performed on 9 urinary exosome (UE) pooled samples by gel based digestion followed by LC-MS/MS, led to the identification of 261 proteins from CTRL subject UE and 186 from RCC patient UE, and demonstrated that most of the identified proteins are membrane associated or cytoplasmic. Moreover, about a half of identified proteins are not shared between RCC and control UE. Starting from these observations, and from the literature, we selected a panel of 10 proteins, whose UE differential content was subjected to immunoblotting validation. Results show for the first time that RCC UE protein content is substantially and reproducibly different from control UE, and that these differences may provide clues for new RCC biomarker discovery. (hide)
EV-METRIC
44% (89th percentile of all experiments on the same sample type)
 Reported
 Not reported
 Not applicable
EV-enriched proteins
Protein analysis: analysis of three or more EV-enriched proteins
non EV-enriched protein
Protein analysis: assessment of a non-EV-enriched protein
qualitative and quantitative analysis
Particle analysis: implementation of both qualitative and quantitative methods
electron microscopy images
Particle analysis: inclusion of a widefield and close-up electron microscopy image
density gradient
Isolation method: density gradient, at least as validation of results attributed to EVs
EV density
Isolation method: reporting of obtained EV density
ultracentrifugation specifics
Isolation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps
antibody specifics
Protein analysis: antibody clone/reference number and dilution
lysate preparation
Protein analysis: lysis buffer composition
Study data
Sample type
Urine
Focus vesicles
exosomes
Isolation protocol
Isolation protocol
  • Gives a short, non-chronological overview of the
    different steps of the isolation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
dUC + Iodixanol-DG (valid.)
Protein markers
EV: CD9/ Flotilin1/ Syntenin/ TSG101/ CD10/ EMMPRIN/ DPEP1/ CP/ MMP9/ Podocalyxin/ CAIX/ DKK4/ AQP1
non-EV: None
Proteomics
yes
EV density (g/ml)
1.100
TEM measurements
41.1+-12.7
Show all info
Study aim
Omics
Sample
Species
Homo sapiens
Sample Type
Urine
Isolation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Equal to or above 150,000 g
Pelleting: time(min)
60
Density gradient
Only used for validation of main results
1
Density medium
Iodixanol
Lowest density fraction
5
Highest density fraction
40
Orientation
Top-down
Speed (g)
100000
Characterization: Protein analysis
Western Blot
Detected EV-associated proteins
CD9/ Flotilin1/ Syntenin/ TSG101/ CD10/ EMMPRIN/ DPEP1/ CP/ MMP9/ Podocalyxin/ CAIX/ DKK4/ AQP1
ELISA
Detected EV-associated proteins
CD10/ EMMPRIN/ DPEP1/ CP/ MMP9/ Podocalyxin/ CAIX/ DKK4/ AQP1
Fluorescent NTA
Characterization: Particle analysis
EM
EM-type
transmission EM
Image type
Wide-field
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