Hi all, I am doing assays with extracellular vesicles (EVs) in my lab and I have read in many articles that relate EVs concentration to protein concentration.
For example, in this article https://doi.org/10.1371/journal.pone.0196080 or in this one doi: 10.3402/jev.v2i0.19861
I want to correlate the protein concentration with the concentration of EVs, but I am still wondering if when measuring the protein concentration by Bradford's Method or BCA, in order to correlate it with the concentration of EVs, do the proteins have to be previously extracted from the EVs or do I just add Bradford's reagent on my sample with intact EVs without any protein extraction?
(I am still going to do NTA analysis, but at the moment I am only concerned about the question I am asking).
Thank you very much in advance!
Hello,
if you are not going to work on proteins that are inside EV or on the EV surface; all this because you want to calculate how much EV do you have (for example if you are gonna treat the cells with EV you should know the concentration) then you can directly use Bradford reagent. All I know is it is hard to isolate proteins from EV because it is hard to disrupt the EV membrane compare to the cell membrane.
I would not try to isolate the protein from EVs if you just want to measure their total protein content. As suggested you should be able to add Bradford reagent directly. If you are concerned then you might not get EV lysis and thus not detect all the EV protein cargo then you could resuspend the EVs in RIPA buffer for a couple of hours on ice, vortex. And then perform Bradford assay on the suspension.
EVs are nano-sized, membrane-bound vesicles that contain several biomolecules, including proteins, lipids, mRNAs, and microRNA. The quantity of protein concentration (using BCA without lysis buffer) in EVs has been found to be correlated with the number of particles in EVs (10.1038/s41598-018-32953-7).
The extracted EV proteins are required for western blot and proteomic analysis. For treatment, measurement of total protein concentration using BCA has been used in several publications.
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