Ammonium sulfate cuts
Dr E. Buxbaum
EB15 at le.ac.uk
Wed Feb 14 08:46:39 EST 1996
Bronwyn Venus <venusb> wrote:
> I need to remove most of the albumin from a lymph sample to enable
> processing through an IEF Rotofor.
> I think the best way to do this is using Ammonium sulfate, ppt albumin
> and use the supernatant.
Remember that you need low conductivity samples for IEF
(isoelectric focussing). Adding a lot of salt is therefore
counterproductive, unless you are prepared to remove the salt later
by gel filtration or repeated dialysis.
Another consideration is what you want to purify. Is there a
significant difference in the solubility of your protein and
albumin in ammonium sulfate? When I consider AS-precipitation
for a separation I usually do a serial experiment with 20, 30,
and so on to 80% saturation (100% saturated AS is about 4.2 M) and
analyse the precipitates and supernatants for my protein. It is
usually possible to get 2-3 fold enrichment with this method.
Any decent book on protein purification should have the details,
check for example the volumes on protein purification in
"Methods in Enzymology" (your library should have this).
For your purpose you may want to consider ethanol or aceton
precipitation instead of AS, they will not increase the
conductivity of your sample.
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