Tris Tricine SDS PAGE Stacking pH?

Dr Engelbert Buxbaum engelbert_buxbaum at hotmail.com
Tue Sep 5 08:09:57 EST 2006


Wolfgang Schechinger wrote:

> Maybe all this reads quite weird to all you Kit(dy)-fellows out here,
> however me just wants (i.e. needs) to get most out of a very limited
> sample material: I'd like to analyse a variety of proteins (10-200 kD
> in the same gel) and their phosphorylation status from human muscle
> biopsies (average 1mg of total protein available, which must be
> sufficient for at least 12 gels), so I need to get the sharpest bands
> available in order to increase the sensitivity of detection.

Then I would use gradient gels rather than switching to a different
buffer system. A 5-15% T gradient Laemli gel should do what you want. If
you do not have a gradient maker you can use a step gradient of a dozen
or so equal spaced steps, in that case it helps to add some 5% glycerol
to the heavy solution in order to increase the density difference
between the solutions.



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