I want to isolate exosomes from serum to do proteomics which need a large volume protein. As known, UC can not get enough protein,some kits like exosomequick can bring some contaminations of serum protein. So does anybody have good methods to isolation serum exosomes.
Thank you very much. Here is a SDS-page of exosomes proteins by different isolation methods. From left, one lane is serum, two is UC, three is 101bio exposure, four is Exoquick, five is marker. Different methods get completely different protein stripe.
I believe that ultracentrifugation is the gold standard in the field. You may need to increase the amount of plasma or serum you start with.
Haihua,
looking at your gel - you have sufficient protein to do proteomics, your UC lane looks excellent. Would it be feasible for you to do a proteomic comparison of the four lanes to answer your question experimentally? That would definitively be the best option.
Alternatively, you could just get one particular region (ideally one that contains a protein or a group of proteins that you are interested in for biological reasons), cut that out from the 4 lanes, do an in-gel digest and compare the results only for that region.
hello there,
you have to centrifuge serum at ~10k rpm so as to remove any cell debris, use the supernatant for Ultra centrifugation (say 45krpm ~2.4 lac g) where you can settle down only exosomes in pellet. now you can resuspend exosomes in appropriate buffer.
Total exosome isolation reagents (from serum) or (from plasma)
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