Need to increase protein precipitation yield w/ TCA precipitation protocol - (Jun/01/2007 )
Whenever I do TCA precipitation w/ the eluted protein solution after immunoprecipitation, I always feel that the precipitated protein yield is too low. Is there any way to increase protein yield with TCA precipitation?
Add first acetone and then TCA. normally it's 30' at -20°.
2h at -80° or more increase the yield.
You may also try lyophilisation for getting higher yield. But often you need dialysis after that as the salts are also lyophilized
if you are not going to apply to a gel and silver stain then you can add a few micrograms of trna to the solution before you add the tca. it will coprecipitate and carry the protein down when you spin it. we used to use this to concentrate samples for coomassie stained gels (we found that we would get a big orange blotch if we silver stained).
I did with TCA first, then acetone. finally I got quite big fat bands in silver stain. And I also can see bands in Coomassie stain. I wondered guys what solution finally dissolve TCA precipitated proteins in? followed by MS?
I am using 10mM tris-cl, pH7.4. What else can get better solubility and good for MS?