hydroxyapatite columns problems

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hydroxyapatite columns problems

Nicolas BAZEILLE
Hi,

I'm trying to purify protein using hydroxypapatite system from bio-rad. The
purification step work well, but the powder is very hard to regenerated.
I don't use any forbidden components in my buffer such EDTA or Tris.
I use different kinds of regeneration pathways (500 mM phosphate, 1M NaOH,
6M guanidium chloride for short time) according to the supplier
informations. However, after regeneration, the flow rate greatly decrease,
and the powder seems to be damaged (it seems to be solubilize while
solution containing phosphate). I don't want to use new product for each
purification  Is there a way to preserve the stability of the material ?
Is somebody find the same problem?

thanks in advance.

M. BAZEILLE Nicolas
Equipe XI
GĂ©notoxicologie et cycle cellulaire - UMR 2027 CNRS/Institut Curie
Centre Universitaire
Bâtiments 110
91405 ORSAY Cedex
01-69-86-71-27  



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Re: hydroxyapatite columns problems

Dr Engelbert Buxbaum
Am 02.02.2009, 12:43 Uhr, schrieb Nicolas BAZEILLE  
<[hidden email]>:

> Hi,
>
> I'm trying to purify protein using hydroxypapatite system from bio-rad.  
> The purification step work well, but the powder is very hard to  
> regenerated.
> I don't use any forbidden components in my buffer such EDTA or Tris.
> I use different kinds of regeneration pathways (500 mM phosphate, 1M  
> NaOH, 6M guanidium chloride for short time)

Hydroxyapatite is a crystalline form of calcium phosphate. It is easier to  
use with a ceramic backbone (BioGel HTP). I am not sure why you use  
conditions like NaOH or guanidinium chloride, with a properly prepared  
sample that should not be necessary.
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