Some people add 1 mg/mL BSA to their sample in order to get the total
protein concentration high enough for precipitation. Of course, then
you're left with lots of BSA in your precipitate too....
On Mar 11, 2010, at 9:27 AM, Nikola Wenta wrote:
Hi Iraz!
I don't have any clue about acetone precipitation of proteins, but
generally, precipitation with saturated Ammonium sulfate solution
provides a good means to concentrate proteins and to get them into
a save state for short-term storage. Unfortunatelly, you would
already need the protein solution to be at > 1 mg/ml in order to
get it to precipitate. Thus, in your case this method doesn't seem
suitable. You could also try to concentrate the protein with
Centricons, but you would rather loose protein to the membrane than
concentrate your solution as it is already too diluted. Why not
loading maximum volume into biggest possible pockets on a gel with
maximum thick spacers? Additionally you could use a acrylamide
percentage that "compresses" your protein band, giving you a better
signals in WB.
Best, Niko
-----Ursprüngliche Nachricht-----
Von: [email protected] im Auftrag von methods-
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Gesendet: Do 11.03.2010 17:03
An: [email protected]
Betreff: Methods Digest, Vol 58, Issue 7
Message: 8
Date: Thu, 11 Mar 2010 16:56:05 +0100
From: "Iraz Toprak Aydin" <[email protected]>
Subject: Protein precipitation - acetone?
To: <[email protected]>
Message-ID: <004401cac133$5b7c0df0$127429...@[email protected]>
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Dear all,
I have to do a western blot, but my protein concentration is very
low, and I
have to run a mini gel. So I was thinking of precipitation the
proteins. I
have never done this before. Does acetone have a bad effect on the
blotting?
Are there any points that I should be careful about?
Thanks in advance...
Iraz Toprak Aydin
EPFL SV ISREC, Station 19
Batiment SV, SV 2540
CH-1015 Lausanne
Switzerland
Tel: +41 21 693 07 36
e-mail: <mailto:[email protected]> [email protected]
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