running sds gel with guanidium

Scott Coutts scott.coutts at med.monash.edu.au
Wed Nov 26 18:25:00 EST 2003

Yes, it wont work, as you have found out! It usually precipitates. Did 
you see it precipitate or become opaque when you loaded your sample in 
the well? Anyway, I assume you're using the guanidinium to keep a 
protein soluble. If not, then this may not apply to your problem (:

But If so, then you need to do some tests to determine whether your 
protein is soluble in SDS, or in urea. If it is, then you need to use 
one of these in preference to guanidinium. Often, this is not possible, 
so in these cases you need to buffer-exchange your sample. You need to 
do this by dialysing against an SDS-PAGE compatible buffer such as urea. 
If you only have a very small volume (too small for easy dialysis using 
dialysis tubing) then you can do drop dialysis. You need to buy an 
ultrafiltration membrane (from millipore for example) that has a pore 
size smaller than your protein (I normally use 10,000 MWCO). Then, put a 
  petri dish or a 50mL tube and carefully place your membrane on top of 
it so that it is dry on one side and floating. Then you need to 
carefully pipette, say, 20 to 50uL drops on to the top of the membrane 
and leave them for a while to dialyse. Then, the hardest part is 
pipetting them back off wihtout sinking the membrane - you need to have 
steady hands (:

Then, they should have exchanged their guanidinium buffer for urea buffer.

Good luck.

ss at nordicbioscience.com wrote:

> I tried to analyse by western-blot protein samples that I have treated with guanidium. But the proteins migrated in a smear, like if all the samples were degraded. Is there a problem to run an SDS page in presence of guanidium?
> http://biowww.net/mynews/tree.php?group_name=bionet_molbio_proteins&begin=0

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