how to remove media components from protein solutions - (Aug/01/2006 )
I am trying to purify a 25 kDa protease that is secreted by an orgainsm into the supernatant of a rich media. Although i have the protein purified, the solution has a medium dark brown tinge that cannot be precipitated. I have tried running it through size fraction columns and also tried dialysing it, but miserably failed. I need to remove these media contaminants before i could make antibodies to this protease. Can anyone help me or suggest a way by which i could remove these media contaminants from the protein solution.
what do you use as size fraction column?
I make use of a P100 resin from BIORAD. We used both fine and medium sized resin, but without any luck.
is it possible to grow your organism up in the rich media, spin it down, and then induce secretion in a minimal media? your total secreted protein yield will probably be lower, but you might have an easier time purifying it.
Have you tried ion exchange column?