FPLC Column Cleaning Protocol?

Donny Wong wong5 at login2.fas.harvard.edu
Tue Jul 21 23:27:28 EST 1998


Dear netters,
I've been having a  horrible time with a mono-S column on the FPLC...the
back pressure is very high, causing the system to shut down very
frequently.  I tried cleaning out the column with a protocol from
Pharmacia (2M salt injection, 1N NaOH inject, 0.5% SDS/0.5 N NaOH), but
the pressure is still quite high when i inject my sample and while washing
the column afterwards.  I managed to inject my entire sample by stopping
the machine every time the pressure got too high, waiting for it to go
down, and then starting up the injection again (I did this for 25mls...and
it shut down pretty much after every ml (at 0.1 ml/min).  finally i am
washing and the pressure is stable, so i may be able to complete the
purification.  so to make a long story short...is there a good protocol
for cleaning this column after i'm done with it so that i (or anyone else
in the lab) wont have to deal with this problem again?  I'm wary of
Pharmacia's protocol...the 0.5% SDS/NaOH solution made the problem even
worse when the SDS precipitated out of solution in the cold cabinet the
system is in.

thanks,
donny wong
wong5 at fas.harvard.edu



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