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separating 64 and 65 kD proteins

Mon Aug 8 21:12:36 EST 1994

I would question Hiranya's suggestion that a gradient gel will enhance the reso
lution of several closely spaced polypeptides.  The principal function of a gra
dient gel is to "resolve a wider size range of proteins than standard uniform c
oncentration gels (from Current Protocols in Molecular Biology, p. 10.2.9)".  I
t stands to reason that resolution of a broad range of polypeptides on a gel wi
ll sacrifice sensitivity within the optimal middle region of the gel.  My exper
iences with the resolution of closely spaced polypeptides are similar to those
described by Rainer, namely a straight percentage gel in which the acrylamide c
oncentration and gel length are optimized for the specific molecular weight cla
ss and gel running conditions are adjusted to minimize 'broad" bands--running t
he gels at low temperature to reduce diffusion and thin gels.

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