PREFRACTIONATION OF PROTEIN SAMPLES PRIOR TO 2-DIMENSIONAL ELECTROPHORESIS

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.