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Native PAGE @ pH < 6.5

Christopher Walentas Bioc cdw1000 at mole.bio.cam.ac.uk
Tue Mar 5 09:00:56 EST 1996

I was wondering if anyone had a buffering system that they could
recommend for the resolution of two proteins using native PAGE at
a pH at or below 6.5?  I've got a mixture of two species which
differ by a series of histidines, and need to utilise their charge
differences (which are only relevant below their pKa), but cannot
go too acidic due their pI is around 4.  I've tried citric acid
at pH 5.5 (100mM) but with not much luck-- the proteins are 100kDa
and hardly made it into the resolving gel by the time the BPBlue
had scooted off the bottom.  I've looked into a number of Practical
Approach-type reference manuals, but for the most part they are too
general and of little or no help.  Any advice in this matter would
be most appreciated.



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