Painful thrombin digestion

ChenHA hzhen at
Sat Mar 19 11:45:25 EST 2005

Belle wrote:
> So, I'm clueless about what to do at this moment. Would anyone have
> any suggestions?

Just a few general suggestions - 

1) If others have got success with thrombin digest without adding
PMSF, then why not try it?

2) Thrombin digest often works better after the GST fusion protein has
been eluted off the beads.

3) I normally do thrombin digest at room temperature, try that unless
you have serious problem with unwanted proteolysis. Check progress by
taking samples after 30 min., 1 hour, etc. 

4) Check the storage condition of the thrombin solution as recommended
by the supplier (I store mine at 4 deg C with no problem) and that the
thrombin is still active.

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