Protein Profiling of Serum Extracellular Vesicles Reveals Qualitative and Quantitative Differences After Differential Ultracentrifugation and ExoQuickTM Isolation
Solid tumor biopsies are the current standard for precision medicine. However, the procedure is invasive and not always feasible. In contrast, liquid biopsies, such as serum enriched for extracellular vesicles (EVs) represent a non-invasive source of cancer biomarkers. In this study, we compared two EV isolation methods in the context of the protein biomarker detection in inflammatory bowel disease (IBD) and colorectal cancer (CRC). Using serum samples of a healthy cohort as well as CRC and IBD patients, EVs were isolated by ultracentrifugation and ExoQuickTM in parallel. EV associated protein profiles were compared by multiplex-fluorescence two-dimensional difference gel electrophoresis (2D-DIGE) and subsequent identification by mass spectrometry. Validation of gelsolin (GSN) was performed using fluorescence-quantitative western blot. 2D-DIGE resolved 936 protein spots in all serum-enriched EVs isolated by ultracentrifugation or ExoQuickTM. Hereof, 93 spots were differently expressed between isolation approaches. Higher levels of GSN in EVs obtained with ExoQuickTM compared to ultracentrifugation were confirmed by western blot (p = 0.0006). Although patient groups were distinguishable after both EV isolation approaches, sample preparation strongly influences EVs' protein profile and thus impacts on inter-study reproducibility, biomarker identification and validation. The results stress the need for strict SOPs in EV research before clinical implementation can be reached.
| Errataetall: | |
|---|---|
| Medienart: |
E-Artikel |
| Erscheinungsjahr: |
2020 |
|---|---|
| Erschienen: |
2020 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:9 |
|---|---|
| Enthalten in: |
Journal of clinical medicine - 9(2020), 5 vom: 12. Mai |
| Sprache: |
Englisch |
|---|
| Beteiligte Personen: |
Gemoll, Timo [VerfasserIn] |
|---|
| Links: |
|---|
| Themen: |
Colorectal cancer |
|---|
| Anmerkungen: |
Date Revised 07.08.2021 published: Electronic ErratumIn: J Clin Med. 2021 Jul 29;10(15):. - PMID 34362233 Citation Status PubMed-not-MEDLINE |
|---|
| doi: |
10.3390/jcm9051429 |
|---|
| funding: |
|
|---|---|
| Förderinstitution / Projekttitel: |
|
| PPN (Katalog-ID): |
NLM309912709 |
|---|
| LEADER | 01000naa a22002652 4500 | ||
|---|---|---|---|
| 001 | NLM309912709 | ||
| 003 | DE-627 | ||
| 005 | 20231225134944.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 231225s2020 xx |||||o 00| ||eng c | ||
| 024 | 7 | |a 10.3390/jcm9051429 |2 doi | |
| 028 | 5 | 2 | |a pubmed24n1033.xml |
| 035 | |a (DE-627)NLM309912709 | ||
| 035 | |a (NLM)32408476 | ||
| 035 | |a (PII)E1429 | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 041 | |a eng | ||
| 100 | 1 | |a Gemoll, Timo |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Protein Profiling of Serum Extracellular Vesicles Reveals Qualitative and Quantitative Differences After Differential Ultracentrifugation and ExoQuickTM Isolation |
| 264 | 1 | |c 2020 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a ƒaComputermedien |b c |2 rdamedia | ||
| 338 | |a ƒa Online-Ressource |b cr |2 rdacarrier | ||
| 500 | |a Date Revised 07.08.2021 | ||
| 500 | |a published: Electronic | ||
| 500 | |a ErratumIn: J Clin Med. 2021 Jul 29;10(15):. - PMID 34362233 | ||
| 500 | |a Citation Status PubMed-not-MEDLINE | ||
| 520 | |a Solid tumor biopsies are the current standard for precision medicine. However, the procedure is invasive and not always feasible. In contrast, liquid biopsies, such as serum enriched for extracellular vesicles (EVs) represent a non-invasive source of cancer biomarkers. In this study, we compared two EV isolation methods in the context of the protein biomarker detection in inflammatory bowel disease (IBD) and colorectal cancer (CRC). Using serum samples of a healthy cohort as well as CRC and IBD patients, EVs were isolated by ultracentrifugation and ExoQuickTM in parallel. EV associated protein profiles were compared by multiplex-fluorescence two-dimensional difference gel electrophoresis (2D-DIGE) and subsequent identification by mass spectrometry. Validation of gelsolin (GSN) was performed using fluorescence-quantitative western blot. 2D-DIGE resolved 936 protein spots in all serum-enriched EVs isolated by ultracentrifugation or ExoQuickTM. Hereof, 93 spots were differently expressed between isolation approaches. Higher levels of GSN in EVs obtained with ExoQuickTM compared to ultracentrifugation were confirmed by western blot (p = 0.0006). Although patient groups were distinguishable after both EV isolation approaches, sample preparation strongly influences EVs' protein profile and thus impacts on inter-study reproducibility, biomarker identification and validation. The results stress the need for strict SOPs in EV research before clinical implementation can be reached | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a colorectal cancer | |
| 650 | 4 | |a extracellular vesicles | |
| 650 | 4 | |a inflammatory bowel disease | |
| 650 | 4 | |a isolation | |
| 650 | 4 | |a proteomics | |
| 700 | 1 | |a Rozanova, Svitlana |e verfasserin |4 aut | |
| 700 | 1 | |a Röder, Christian |e verfasserin |4 aut | |
| 700 | 1 | |a Hartwig, Sonja |e verfasserin |4 aut | |
| 700 | 1 | |a Kalthoff, Holger |e verfasserin |4 aut | |
| 700 | 1 | |a Lehr, Stefan |e verfasserin |4 aut | |
| 700 | 1 | |a ElSharawy, Abdou |e verfasserin |4 aut | |
| 700 | 1 | |a Habermann, Jens K |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t Journal of clinical medicine |d 2012 |g 9(2020), 5 vom: 12. Mai |w (DE-627)NLM230666310 |x 2077-0383 |7 nnns |
| 773 | 1 | 8 | |g volume:9 |g year:2020 |g number:5 |g day:12 |g month:05 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.3390/jcm9051429 |3 Volltext |
| 912 | |a GBV_USEFLAG_A | ||
| 912 | |a GBV_NLM | ||
| 951 | |a AR | ||
| 952 | |d 9 |j 2020 |e 5 |b 12 |c 05 | ||