Problem with Pierce GST spin purification column
Posted 11 June 2011 - 10:00 AM
Posted 23 June 2011 - 12:29 AM
Posted 07 July 2011 - 01:14 PM
can you tell me which company's sepharose beads do you use?
Posted 15 July 2011 - 07:09 AM
Edited by mdfenko, 15 July 2011 - 07:10 AM.
genius does what it must
i do what i get paid to do
Posted 15 July 2011 - 03:09 PM
Posted 17 July 2011 - 05:51 PM
Thanks! i jus figured that out!
Posted 17 July 2011 - 06:18 PM
Yes the column is prepacked. The protocol says that the lysate should be added to the column and incubated in an end-to-end rotator. However the resin gets dislodged in the subsequent centrifugation steps and it becomes difficult as the resin gets stuck in the middle of the column. When I just add more wash buffer and centrifuge to make the resin settle down I think some of glutathione beads get removed in the flow through and hence I lose protein in the flow through. I have tried avoiding incubating in the rotator and just let the lysate pass through (gravity flow) but lysate does not pass through and gets stuck. As soon as I centrifuge the column the resin gets dislodged.