Hello,
I am doing a western blot on human T-cell lysates. I already made a protein lysate using sonication and SDS/glycerol/TrisHCL/CPI buffer. Now, my concentrations are too low. I was wondering what concentration method works best for these samples?
People are generaly using combination of aceton and deoxycholate.
But I have very good experience with this method: Article A method for the quantitative recovery of protein in dilute ...
It can handle glycerol and detergents in the protein solution.
In this case , Acetone precipitation works quite well , It is very simple , Just add 3 volumes of Ice cold acetone to your lysate , keep it in -20 from 20-60min , afterwards , you will see white precipitate of your proteins , Pelleted down and resuspended in smaller volume and measure protein conc.
Good Luck!
You may use spin membrane microfilter with a cut off of 10 kDa or 5 kDa.
@ Shamsher Singh Kanwar Well, there is SDS in the lysate so I wonder if unfolded smaller proteins would not go through the membrane microfilter.
Do you think your lysate contain proteins smaller than 5 kDa molecular mass?
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