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Ammonium bicarbonate in IE chromatography

Ammonium bicarbonate in IE chromatography - Protein Forum

Ammonium bicarbonate in IE chromatography - Protein Forum


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  #1  
Old 08-31-2006, 10:26 AM
pawel.stocki@ncl.ac.uk
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Default Ammonium bicarbonate in IE chromatography



Hi,
Does anyone have some experience in using ammonium bicarbonate buffer
for gradient elution from Q-Sepharose or other IE? I would like to use
it instead of NaCl gradient. The sample will be subjected for further
MS analysis that is why I would like to use volatile salt. I'll
appreciate any comments.

Thanks,
Pawel

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  #2  
Old 09-01-2006, 02:05 PM
Rhodobacter
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Default Ammonium bicarbonate in IE chromatography

Hi pawel,

its better to dialyse or ultrafiltrate your sample after the IE
chromatography!
Alternativly you can use a desalting column or ZIPTIPs (e.g. Qiagen).

For most samples you need strong Ions in the IE chromatography. May be that
the carbonate degas as CO2 and you will get some bubbles in your column.

Good Luck

CZ

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  #3  
Old 09-04-2006, 03:09 AM
Dr Engelbert Buxbaum
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Default Ammonium bicarbonate in IE chromatography

Rhodobacter wrote:


All this works only for reasonably large sample molecules. Purification
of small molecules by IEC equires volatile buffers that can be removed
by lyophilisation.


Probably not on the column, because of the high pressure there. But
storage of the buffer is not possible, and buffer decomposition can
occur in the fractions, leading to a shift in pH. This can happen also
during lyophilisation, one reason why I have stopped to use acetate or
formiate based buffers, even though they are often recommended (the acid
goes of slower than the ammonia or trimethylammonia and makes the sample
strongly acidic).
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