Skip to content
2000
  1. Home
  2. A-Z Publications
  3. Recent Patents on Biotechnology
  4. Fast Track Articles
  5. Fast Track Article
image of Comparison of the Characteristics of Circulating Small Extracellular Vesicles Isolated by Ultracentrifugation and a Commercial Kit

Abstract

Introduction

The market offers a wide range of extracellular vesicles (EVs) isolation products, but their lack of standardization is a concern. Therefore, it is important to carefully assess the quality of the EVs obtained using these products. In this study, we compared the EXOCIB kit with the ultracentrifuge method, which is considered the gold standard for small EV isolation.

Methods

After overnight fasting, small plasma EVs were extracted from four individuals using both the ultracentrifuge and the EXOCIB kit methods. The pooled EVs were then compared for the presence of the cluster of differentiation 63 (CD63) protein using the western blot analysis, and their size and zeta potential were performed by Dynamic Light Scattering (DLS). In addition, the size and morphology of small EVs were determined by using the Transmission Electron Microscopy (TEM) technique.

Results

An average hydrodynamic size of 135.7 nm and a zeta potential of -6.33 Mv at 25°C was found for small EVs isolated by the ultracentrifuge, whereas the kit method resulted in small EVs with a hydrodynamic size of 102.8 nm and a zeta potential of -0.907. Notably, the size of the particles in the kit samples was smaller compared to those obtained through the ultracentrifuge (P < 0.001). The western blot method confirmed the expression of CD63 in both methods, so the ultracentrifuge yielded small EVs with a higher level of purity compared to the kit-based approach (P = 0.036).

Conclusion

The DLS findings revealed the existence of vesicles within the appropriate size range for small EVs like exosomes in both isolation techniques. The results of the western blot analysis, in conjunction with DLS, displayed that the ultracentrifuge method extracted small EVs with a greater degree of purity than the kit-based approach.

© 2024 Bentham Science Publishers
Loading

Article metrics loading...

/content/journals/biot/10.2174/0118722083325164241015103217
2024-10-25
2025-06-17

  • From This Site

    /content/journals/biot/10.2174/0118722083325164241015103217
    dcterms_title,dcterms_subject,pub_keyword
    -contentType:Contributor -contentType:Concept -contentType:Institution
    10
    5
Loading full text...

Full text loading...

References

  1. Welsh J.A. Goberdhan D.C.I. O’Driscoll L. Mini-mal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches. J. Extracell. Vesicles 2024 13 2 e12404 10.1002/jev2.12404 38326288
    [Google Scholar]
  2. Sheta M. Taha E.A. Lu Y. Eguchi T. Extracellular ves-icles: New classification and tumor immunosuppres-sion. Biology (Basel) 2023 12 1 110 10.3390/biology12010110 36671802
    [Google Scholar]
  3. Xu T. Yu X. Xu K. Comparison of the ability of exosomes and ectosomes derived from adipose-derived stromal cells to promote cartilage regenera-tion in a rat osteochondral defect model. Stem Cell Res. Ther. 2024 15 1 18 10.1186/s13287‑024‑03632‑4 38229196
    [Google Scholar]
  4. Meldolesi J. Exosomes and ectosomes in intercellular communication. Curr. Biol. 2018 28 8 R435 R444 10.1016/j.cub.2018.01.059 29689228
    [Google Scholar]
  5. Afrisham R. Sadegh-Nejadi S. Meshkani R. Emamg-holipour S. Bagherieh M. Paknejad M. Anti-inflammatory effects of plasma circulating exosomes obtained from normal-weight and obese subjects on hepatocytes. Endocr. Metab. Immune Disord. Drug Targets 2021 21 3 478 484 10.2174/1871530320666200505121426
    [Google Scholar]
  6. Sadegh-Nejadi S. Afrisham R. Emamgholipour S. Influence of plasma circulating exosomes ob-tained from obese women on tumorigenesis and ta-moxifen resistance in MCF ‐7 cells. IUBMB Life 2020 72 9 1930 1940 10.1002/iub.2305 32542981
    [Google Scholar]
  7. Afrisham R. Sadegh-Nejadi S. Meshkani R. Emamg-holipour S. Paknejad M. Effect of circulating exo-somes derived from normal-weight and obese women on gluconeogenesis, glycogenesis, lipogenesis and se-cretion of FGF21 and fetuin A in HepG2 cells. Diabetol. Metab. Syndr. 2020 12 1 32 10.1186/s13098‑020‑00540‑4 32322309
    [Google Scholar]
  8. He G. Peng X. Wei S. Exosomes in the hypoxic TME: From release, uptake and biofunctions to clini-cal applications. Mol. Cancer 2022 21 1 19 10.1186/s12943‑021‑01440‑5 35039054
    [Google Scholar]
  9. Kugeratski F.G. Kalluri R. Exosomes as mediators of immune regulation and immunotherapy in cancer. FEBS J. 2021 288 1 10 35 10.1111/febs.15558 32910536
    [Google Scholar]
  10. Saad M.H. Badierah R. Redwan E.M. El-Fakharany E.M. A comprehensive insight into the role of exo-somes in viral infection: Dual faces bearing different functions. Pharmaceutics 2021 13 9 1405 10.3390/pharmaceutics13091405 34575480
    [Google Scholar]
  11. Xiao L. Hareendran S. Loh Y.P. Function of exo-somes in neurological disorders and brain tumors. Ex-tracell Vesicles Circ Nucl Acids 2021 2 55 79 10.20517/evcna.2021.04
    [Google Scholar]
  12. Martins T.S. Vaz M. Henriques A.G. A review on com-parative studies addressing exosome isolation meth-ods from body fluids. Anal. Bioanal. Chem. 2023 415 7 1239 1263 10.1007/s00216‑022‑04174‑5 35838769
    [Google Scholar]
  13. Kurian T.K. Banik S. Gopal D. Chakrabarti S. Ma-zumder N. Elucidating methods for isolation and quantification of exosomes: A review Mol. Biotechnol. 2021 63 4 249 266 10.1007/s12033‑021‑00300‑3 33492613
    [Google Scholar]
  14. Li W.J. Chen H. Tong M.L. Niu J.J. Zhu X.Z. Lin L.R. Comparison of the yield and purity of plasma exo-somes extracted by ultracentrifugation, precipitation, and membrane-based approaches. Open Chem. 2022 20 1 182 191 10.1515/chem‑2022‑0139
    [Google Scholar]
  15. Alegre E. Zubiri L. Perez-Gracia J.L. Circulating melanoma exosomes as diagnostic and prognosis bi-omarkers. Clin. Chim. Acta 2016 454 28 32 10.1016/j.cca.2015.12.031 26724367
    [Google Scholar]
  16. Caradec J. Kharmate G. Hosseini-Beheshti E. Ado-mat H. Gleave M. Guns E. Reproducibility and effi-ciency of serum-derived exosome extraction meth-ods. Clin. Biochem. 2014 47 13-14 1286 1292 10.1016/j.clinbiochem.2014.06.011 24956264
    [Google Scholar]
  17. Coughlan C. Bruce K.D. Burgy O. Exosome iso-lation by ultracentrifugation and precipitation and techniques for downstream analyses. Curr. Protoc. Cell Biol. 2020 88 1 e110 10.1002/cpcb.110 32633898
    [Google Scholar]
  18. Helwa I. Cai J. Drewry M.D. A comparative study of serum exosome isolation using differential ultracentrifugation and three commercial reagents. PLoS One 2017 12 1 e0170628 10.1371/journal.pone.0170628 28114422
    [Google Scholar]
  19. Gemoll T. Strohkamp S. Rozanova S. Protein profiling of serum extracellular vesicles reveals quali-tative and quantitative differences after differential ultracentrifugation and ExoQuick™ isolation. J. Clin. Med. 2020 9 5 1429 10.3390/jcm9051429 32408476
    [Google Scholar]
  20. Farrokhi V. Afrisham R. Soleimani M. The ef-fect of circulating exosomes obtained from young and old individuals on the aging related hTERT and P16 expression in hematopoietic stem cells. Nat. Prod. J. 2024 14 8 e140324227998 10.2174/0122103155285692240301052306
    [Google Scholar]
  21. Moradi R. Afrisham R. Kashanikhatib Z. Mousavi S.H. Soleimani M. Alizadeh S. The comparative effect of plasma exosomes of young and old people on the expression of BCL-2 and BAX genes in hematopoiet-ic stem cells. Indian J. Hematol. Blood Transfus. 2024 2024 1 8 10.1007/s12288‑024‑01779‑x
    [Google Scholar]
  22. Aziz M.A. Seo B. Hussaini H.M. Hibma M. Rich A.M. Comparing two methods for the isolation of exo-somes. J. Nucleic Acids 2022 2022 1 1 6 10.1155/2022/8648373 36329709
    [Google Scholar]
  23. Macías M. Rebmann V. Mateos B. Comparison of six commercial serum exosome isolation methods suitable for clinical laboratories. Effect in cytokine analysis. Clin. Chem. Lab. Med. 2019 57 10 1539 1545 10.1515/cclm‑2018‑1297 30990781
    [Google Scholar]
  24. Patel G.K. Khan M.A. Zubair H. Comparative analysis of exosome isolation methods using culture supernatant for optimum yield, purity and down-stream applications. Sci. Rep. 2019 9 1 5335 10.1038/s41598‑019‑41800‑2 30926864
    [Google Scholar]
  25. Skottvoll F.S. Berg H.E. Bjørseth K. Ultracentrif-ugation versus kit exosome isolation: NanoLC-MS and other tools reveal similar performance bi-omarkers, but also contaminations. Future Sci. OA 2018 5 1 FSO359 10.4155/fsoa‑2018‑0088 30652024
    [Google Scholar]
  26. Kalarikkal S.P. Prasad D. Kasiappan R. Chaudhari S.R. Sundaram G.M. A cost-effective polyethylene glycol-based method for the isolation of functional edible nanoparticles from ginger rhizomes. Sci. Rep. 2020 10 1 4456 10.1038/s41598‑020‑61358‑8 32157137
    [Google Scholar]
  27. Jia L. Li B. Fang C. Extracellular vesicles of mesenchymal stem cells are more effectively ac-cessed through polyethylene glycol-based precipita-tion than by ultracentrifugation. Stem Cells Int. 2022 2022 1 1 12 10.1155/2022/3577015 36110890
    [Google Scholar]
  28. Yakubovich E.I. Polischouk A.G. Evtushenko V.I. Prin-ciples and problems of exosome isolation from bio-logical fluids. Biochemistry (Moscow). Biochem. Suppl. Ser. A: Membr. Cell Biol. 2022 16 2 115 126 10.1134/S1990747822030096 35730027
    [Google Scholar]
  29. Lozano-Andrés E. Enciso-Martinez A. Gijsbers A. Physical association of low density lipoprotein particles and extracellular vesicles unveiled by single particle analysis. J. Extracell. Vesicles 2023 12 11 12376 10.1002/jev2.12376 37942918
    [Google Scholar]
  30. Simonsen J.B. What are we looking at? Extracellular vesicles, lipoproteins, or both? Circ. Res. 2017 121 8 920 922 10.1161/CIRCRESAHA.117.311767 28963190
    [Google Scholar]
/content/journals/biot/10.2174/0118722083325164241015103217
Loading
Comparison of the Characteristics of Circulating Small Extracellular Vesicles Isolated by Ultracentrifugation and a Commercial Kit
Bentham Science Publishers ; https://doi.org/10.2174/0118722083325164241015103217
/content/journals/biot/10.2174/0118722083325164241015103217
/content/journals/biot/10.2174/0118722083325164241015103217
Loading

Data & Media loading...


  • Article Type:     Research Article
Keywords: ultracentrifugation ; extracellular vesicles ; extracellular vesicles extraction kit ; extracellular vesicles isolation ; Plasma ; Commercial kit ; Exosome

Most Cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error
Please enter a valid_number test