Acid Native PAGE
randall.cameron2 at gte.net
Tue Nov 7 15:04:07 EST 2000
Check Thomas, JM and Hodes, ME. 1981. A new discontinuous buffer system for
electrophoresis of cationic proteins at near-neutral pH. Anal. Biochem.
118:194-196. If you try it be sure to use the MOPS free acid, not the salt.
It works well for slab gels as well as tube gels.
lou <sheridal at mrc-lmb.cam.ac.uk> wrote in message
news:3A083773.DA62FBF9 at mrc-lmb.cam.ac.uk...
> I have two proteins, one with a pI of 8.8 (Mw=50kDa) and the other with
> a pI of 8.3 (Mw=90kDa) I am trying to make a complex of the two proteins
> (which should be irreversible) and would like to see this complex
> formation, preferably on a native gel.
> I have attempted to run an acid native PAGE,. I have made my gels (25mM
> Tris Glycine, 12% Acrylamide) and the buffer is Tris (25mM) Alanine,
> pH7.4. The buffer that the samples are in vary but all are 25mM buffer
> with 100mM NaCl. The loading dye contains glycerol and methyl green.
> The problem that I am having is that the samples do not run very far and
> the bands are quite broad. I have heard that lowering the salt
> concentration of the samples can help with resolution, but by how much.
> Also does anyone know if the pH that I have tried is low enough?? Should
> I just try another gel at lower pH?? Is there any other way in which
> people might be able to advise me??
> I hope that someone can help
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