Protein precipitation

Frank O. Fackelmayer Frank.Fackelmayer at uni-konstanz.de
Thu Sep 23 09:09:14 EST 1999



Peter wrote:

> In article <37E88028.1ED955C5 at uni-konstanz.de>,
> Frank.Fackelmayer at uni-konstanz.de wrote:
>
> > What I routinely use for protein precipitation is a method described by
> > Wessel &  Fluegge (1984), Anal. Biochem. 138:141-143,
>
> I thought that the Wessel Fluegge method was an "extraction" rather than a
> "precipitaion".

No. In fact, what you do is adding methanol and chloroform to "extract" the
protein from a sample solution into the interphase and organic phase (quite
like in phenol/chloroform extraction of DNA, but using the other phase for
furpher processing), then removing the aquous phase, and precipitating the
protein by adding more methanol. You end up with a protein pellet that you
air-dry and redissolve in SDS-PAGE sample buffer. It is sometimes hard to
redissolve the pellet, especially if the pellet is very big (milligram
amounts of total protein). In that case, sonication in SDS-PAGE sample buffer
to break down the pellet before boiling the sample usually helps. Best
results are obtained if you spin your sample after boiling to remove all
residual solids, and use the supernatant only.

Frank




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