Chromatography Forum

Hi,
I am using Amersham's pre-packed Superdex 200 column and their FPLC system.The problem is, whenever I run the column, a gap begins to develop at the top and the column seems to start compressing.When I run the column inverted,the matrix is restored to normalcy.I have washed the column with 20%ethanol to remove any particulate matter stuck inside,but didn't see anything come out in the UV.This keeps happening and leads to an increasingly high back pressure.
Any suggestions please?
Saraswathi

I have used the same column and it is very important that you don't introduce any air in the column. I usually pump without the column for a while to be sure that there is no bubbles coming out, then I run the column backwards for a minute (to eliminate the air in the inlet capillary).

I could also be that the pressure is too high, which that makes the column collapse. Either the flow is too high or you have injected particles.

hope it helps!

thanx.Will try.