Hi!
Does someone know how to clean lipoproteins from EVs? (from human plasma)
We are trying to use iodixanol sucrose gradient (Optiprep), but we don't see EVs markers on the western blot after that process. We did firstly the SEC and then took 500ul to Optiprep ultracentrifugation and also we tried to go directly to Optiprep with 1ml plasma. Both experiments don't work.
What could be the problem?
Do you know about other methods for cleaning the lipoproteins?
Thank you!
Linoy.
Seemed that you have applied the gold standard methods. Orthogonal approaches SEC followed by density gradient either by optiprep or sucrose cushion should be efficient according to literature...If you select the accurate markers and western setup (antibodies and protocols) it must be conclusive. It is hard to say something about inefficient result without knowing the details of your sub-procedural steps. But if you are not targeting functionality but the composition of EV, you may try the affinity isolation and enrichment by magnetic bead coupled with antibodies/lectins/peptides or aptamers...You may get rid of co-eluted lipoproteins and enrich your EVs if you are targeting some special ones. For whole EV profile or investigating the functions of any EVs it is not amenable unfortunately.
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