I extract blood Exosomes by Exoquick kit of SBI , but at the same time enrich a lot of serum lipoproteins, about 1mg in 1ml serum. Lipoprotein volume is very large, serious block 2D-DIGE gel, how can I remove the impact of serum lipoproteins. I will be thankful for your help.The 2D-DIGE gel need 1mg total protein a sample ,so as EXO-FLOW is very expensive.
Dear Haihua,
With exoquick you are not likely to be able to get rid of the lipoprotein contamination. I think it will be much easier for your, much cheaper and less time-consuming of you reconsider the kit that you are using for extraction of your exosomes, instead of using secondary methods for cleaning the samples. There are kits commercially available from the brand pureexo that are just perfect for your case. If you are interested in more details you may write me an e-mail at [email protected]
All Best,
Vasiliy
Thank u very much,I have connected with the manager of prueexo in china,they also feel confused about the working principle of this product.Do you know how it works,can it exclude other microparticles'contamination (eg.lipoprotein).I would very appreciate your help.
consider the Total exosome isolation reagent- its superior for recovery from serum. Keep the vol to minimum- ideally 100 uL serum.
One way to minimize the effects of lipids is to filter the plasma through an 80 um filter, before proceeding with the exosomal isolation.
Hi Haihua,
Serum/blood is a very rich starting material in both exosomes and other common protein contaminants. For your specific concern, I would recommend exploring ExoQuick PLUS. It allows for the enrichment of exosomes from many of the common protein contaminants that can typically be found in serum isolations. Yields should be sufficient for your application and may yield you better downstream results for proteomic/biomarker discoveries.
Best of luck
Hi Haihua,
A combination of size- and density based methods (e.g. size-exclusion chromatography and density gradient centrifugation) can be used to largely separate EVs and lipoprotein particles.
It is possible to use precipitated material from plasma as a starting point:
Article A novel three step protocol to isolate extracellular vesicle...
Other studies that may be useful:
Article Detailed analysis of the plasma extracellular vesicle proteo...
and
Article Y-RNA subtype ratios in plasma extracellular vesicles are ce...
Also discussed in this WebEVTalk by prof. An Hendrix:
https://youtu.be/rIdUUFsEiqo?t=1281
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