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question total exosome isolation protocol( from cell culture media)

according to  total exosome isolation protocol ( from  cell culture media) , before add 0.5 volume kit, Centrifuge the  cell media at   2000  × g   for 30 minutes to remove cells and  debris. My question is the protocol didn't remove microvesicles by 10000g. So the final pellet by kit does not only contain exosomes but also microvesicles.
asked Dec 8, 2013 in ExosomesTalk by anonymous
Plz confirm the above Question?238-0

2 Answers

according to our Nanosight and EM analysis the exosome preps are pretty clean, especially isolated from cell media. Very few particles >300 nm are recovered; the majority of vesicles are in a typical exosome range 30-150 nm.
answered Dec 9, 2013 by Vlassosv
We have used Dynabeads coated with anti-CD63, CD81 or CD9 to pull out vesicles from pre-enriched preparations using Total Exosome Isolation kit and prepare them for electron microscopy. The size distribution of the vesicle located on the surface of the magnetic beads look very similar and small in size (<100nm) and also similar to exosomes prepared by ultracentrifugation.

For exosome workflow see:
- Oksvold et al Clinical Therapeutics 2014 Vol 36 no 6
- Oksvold et al Methods Mol Biol 2015, 1218, 465-81.
- Zeringer et al World J Methodol 03/2013; 26(3(1))
answered Dec 18, 2014 by ketil.pedersen