Thousands of proteins may be visualised on a two-dimensional (2-D) gel
, but only hundreds are present at levels sufficient for chemical anal
ysis. Therefore, prefractionation of protein samples prior to 2-D poly
acrylamide gel electrophoresis (PAGE) will be important for the invest
igation of proteins that are present at sub-picogram levels in physiol
ogical samples. We describe an approach to prefractionate protein samp
les prior to 2-D PAGE using the Gradiflow, which is a new (preparative
) electrokinetic membrane apparatus designed to fractionate proteins i
n a number of different ways. We have fractionated human serum under n
onreducing conditions using the 'reflux' mode, in which proteins are f
ractionated according to their relative mobility under controlled elec
trophoretic conditions, where the current is periodically reversed. We
describe how fractionation occurs and present examples of enrichment
of specific proteins.