Cannot bump GST-proteins off the GSH-beads

[Richard P Grant]

ct133 at columbia.edu wrote:
} Hi,

} I need some help with how to bump off my GST-fusion proteins from the
} GSH-beads. I've tried to bump off with 5, 10 and 15 mM of GSH in 50 mM
} Tris pH 7.5. I've gone up to 20 mM and still couldn't bump the
} proteins off. Is there any trick I can do?

No guarantees, but check the pH of your elution buffer.  It needs to be
around 7.6 to work effectively - and as you increase [GSH], you will need
more buffering capacity.

You could also try at different temperatures - try 5 minutes at room
temperature (which is about 30C here, so maybe a bit lower (-; ) or 30
mins at 4C if the pH is fine.

HTH

R

--
Richard P. Grant  MA  DPhil          University of Oxford
Nuffield Department Obstetrics and Gynaecology      FFPGP
rgrant'at'molbiol.ox.ac.uk            http://www.molbiol.ox.ac.uk/~rgrant
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