FPLC Column Cleaning Protocol?

Dr E. Buxbaum EB15 at le.ac.uk
Mon Jul 27 18:17:16 EST 1998


Donny Wong wrote:
> is there a good protocol for cleaning this column after i'm done with 

This depends somewhat on what you have been loading onto it. If the
problem is precipitated protein, an urea gradient might help (and if it
does, it may be worthwhile to have urea in future samples too). With
lipid you may use a gradient 0-100% Methanol, followed by a
Methanol-Chloroform gradient and the same way back. If you have
particulates in your sample, you may want to think about appropriate
sample preparation in the first place (a few minutes in a table top
centrifuge can spare you a lot of grief). All buffers of course should
be vacuum filtered over a 0.22 um filter both to remove air and
particulates (one would not believe how much sot there is in analytical
grade salts). 

With Pharmacia columns you should be able to remove the piston from the
top and change the net at the end of it, if that has become clogged. Be
carefull though not to introduce air bubbles in the process (ie have the
pump running while you remove the piston, so that the void fills with
buffer instead of air). 

As I said, it depends on what the problem is.



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