Help! A Clogged HIC Cartridge (Bio-Rad)

Anonymous ANONYMOUS.USER at Sunderland.AC.UK
Thu Jul 13 09:08:23 EST 1995


In article <3tsrev$s6b at charm.magnus.acs.ohio-state.edu>, 
mhsieh at magnus.acs.ohio-state.edu (Ming-Ching Hsieh) says:
>
>Hi Dear Netters:
>Can anyone tell me how to recover a clogged HIC cartridge, which is an 
>Econo-Pac Methyl HIC cartridge made by Bio-Rad? I'm using it to purify a 
plant 
>apoplastic protein. I broght the infiltration solution to 2.4 M ammonium 
>sulfate and applied it to the cartridge. The proteins were eluted with a linear
>gradient of ammonium sulfate from 2.4 M to zero. The specific enzyme came 
out 
>at the end of gradient. In the beginning, the flow rate was about 1 ml/min. 
>However, after several uses, the flow rate was reduced to 1 ml/hr. I did clean 
>the cartridge as suggested by the manufacturer, using 100mM NaOH, 20% 
glycerol,
>and deionized water to wash. And I even washed it with 0.5M NaOH and 70% 
>ethanol. They all didn't work. Please tell me what else I can do. Thanks a lot 
>in advance.
>
it sounds like you have blocked the column frit or some things precipertated 
out in the column.
if its the first you could try back flushing the column at a low flow rate say 0.5 
ml/min  for a couple of minutes, but take care nbot to over do this so as disturb 
the column matrix.
if something ppt out then you really have got problems,  you've used alkali to 
wash through the column, would it stand a wash with dilute acid? and then 
deionized water before trying again with the normal reckomended wash cycle.

hope this is of some help.
len bell.
sunderland.



More information about the Proteins mailing list