We have tried TSG101, Actin, LAMP1 as well as CD9. Unfortunately we don't always get the best blots from these antibodies. Can anyone suggest a best control and from where they purchase their antibodies from? Thanks
I just purchased CD9 from System Biosciences and it is VERY strong. They suggest 1:1000 and I think I could do 1:10000, so factor that into their price.
Hi, In terms of western blotting detection antibodies we have 3 different Abs (anti-CD9, anti-CD63 and anti-CD81) that works well on exosomes that have been pre-enriched either by ultracentrifugation or by precipiation and prepared for electrophoresis (non-red conditions) and western blotting.
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ketil
anyone try Alix? just tried myself and didn't have success with it, the other protein I was blotting for came super strong, would've been nice to verify it was in fact exosomes. Looks like I'll look into purchasing CD9
I guess CD9 and CD81 are the go to!
http://www.journalofextracellularvesicles.net/index.php/jev/article/view/20920
the expression level of the different markers depends of course on which cell type the exosomes originate from - as we saw that exosomes derived from B-cell lymphoma cell lines (Ramos, SUDHL-4 and 6 and Ros50 did not seem to contain any CD9 (netiher did the host cells) (Oksvold et al 2014)
I agree with Christian and Ketil.
I'm new to the field but I believe that the exosomes markers can't be used as 'loading controls'. Their expression and expression levels depend on many factors and can't be used as GAPDH/tubulin/actin as we would do with cell-derived proteins.
Thanks for all these precious comments!
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