Hello,
I have a question about exosome-depleted FBS.
I cannot perform ultracentrifugation at 12000 g for 18 hours. Is ultracentrifugation at 100000 g for 1 hour efficient in obtaining exosome-free FBS?
I appreciate your help.
Hello Farnaz Moghadam
Ultracentrifugation at 100000 g for 1 hour won’t be efficient in obtaining exosome-free FBS. Such short protocol will only help to reduce the EVs in FBS to a little extent, and culture medium supplemented with such type of FBS will still have significant effects on cell physiology and phenotype.
I have attached a paper wherein the investigators compared 18 hours ultracentrifugation protocol with a 1.5-hour protocol. The 18-hours ultracentrifugation protocol eliminates approximately 95% of RNA-containing FBS EVs, whereas a 1.5-hour protocol is insufficient.
You may want to refer to the paper attached below.
Article Importance of exosome depletion protocols to eliminate funct...
There are other cost-effective strategies which you could use for the depletion of endogenous EVs from FBS. You could use polymer precipitation and ultrafiltration methods to deplete EVs from FBS samples. These methods are faster and less labor-intensive in comparison to the ultracentrifugation method.
You may want to refer to the article attached below for these methods.
https://jcmr.um.ac.ir/article_29763_9f4cca69c5f62674bd0e7220bdf1873d.pdf
There are also FBS exosome depletion kits which you could make use of. I have attached a link below for one such kit.
https://norgenbiotek.com/sites/default/files/resources/PI61100-3%20FBS%20Exosome%20Depletion%20Kits%20-%20Slurry%20Format.pdf
The kit does not require ultracentrifugation, any special instrumentation, precipitation reagents or any protease treatments. Also, the depletion process is an inexpensive method for depletion of your own FBS, as compared to the current ready-to-use exosome depleted media available commercially.
Best.
Dear Malcolm Nobre ,
I appreciate your recommendations. They were helpful. Thank you for generously sharing your time to help me.
there are centrifuge tubes with filters attached to it, it is effective for exosome isolation rather wasting much time in ultracentrifugation.
dear Jaishree Sankaranarayanan ,
Thank you for your answer. What a great method!
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