I want to remove/inactivate all proteins and peptides from my solution. I was thinking of using proteinase K to cut them all. Any idea of what concentration/time I should use, or any other method which could allow me to do so?
If its a serum-free media then you can do acetone or TCA precipitation. If it has serum then you are out of luck.
You can precipitate all your protein by adding concentrated perchloric acid in your sample, to a final concentration of 5% (use 65% PCA so that you don't dilute your sample too much). You then have to vortex the samples, spin them down to get rid of the protein pellet, collect the supernatant and neutralise it with concentrated KOH (each sample has to be neutralised/pH checked individually). That will create a precipitate of KCl, that you'll want to get rid of by centrifugation again.
Another method, if whatever you're interested in can take a high temperature, is to incubate your samples at 85C for 10min, then spin your samples down. Your denatured proteins will be in the pellet - no need to play with acid/base, much more convenient than above, again if your product of interest can stand the heat.
Thanks all.
The thing is that I want to use the media again to treat other cells after the treatment. Do you think I can still use it after PCA precipitation?
Concerning the heat, I tried it, but I know that some peptides may resist this treatment, so I want to back it up with another method.
If you are going to reuse the media, its probably easiest to use spin filters with varying MW cutoffs.
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