Desalting

[Marcus, Dr J.]

> We wish to purify an enzyme using ion exchange from serum-free conditioned
> medium.  The NaCl concentration is approximately 100 mM.  We would like to
> desalt this medium prior to binding to DEAE-sephadex.  We have tried to
> desalt using buffer exchange (Centri-Prep), however, the enzyme loses its
> activity after desalting by this approach, even if NaCl concentrations are
> raised back to physiological levels!
> 
> Are there any other techniques available that may be used to desalt the
> conditioned medium?
> 
What about straight forward dialysis?  Another thought is 
to try loading your material directly on to your 
ion-exchange column; it just might stick with 100mM NaCl.  
If it does stick, then you no longer have to worry about 
desalting.  If your volumes are not too large you might also be 
able to dilute your sample with an equal volume of buffer 
with no NaCl, thus lowering the salt concentration
and allowing your protein to bind to the column.  

Hoping that helps,
John




_________________________________________________________
John Marcus            Marcus at tpp.uq.edu.au (Dr J.Marcus)
Cooperative Research Centre for Tropical Plant Pathology
5th Level John Hines Building
University of Queensland
St. Lucia, QLD 4072
AUSTRALIA
Fax: 61-7-365-4771
Phone: 61-7-365-4764