(1) Twelve solutions of 0.1 M boric acid and varying glycerol concentration were used to vary pH in a twelve-compartment commercial recycling isoelectric focusing (RIEF) system.
(2) Continuous flow zone electrophoresis (CFE) and recycling isoelectric focusing (RIEF) are two of the alternative formats for fluid phase preparative isolation of biological products in liquid separation media.
(3) RIEF clearly separated each protein with no detectable contamination.
(4) These data demonstrate the superiority of RIEF over CFE for resolution of protein mixtures having only minor charge differences.
(5) Gel filtration chromatography removed most of the remaining, higher molecular weight contaminants of the RIEF-purified enzyme.
(6) Mixtures of 1) cytochrome c, myoglobin, and ovalbumin or 2) beta-lactoglobulin and ovalbumin were used to evaluate the resolving capabilities of CFE and RIEF.
(7) The ground-based McDonnell Douglas CFE and RIEF were compared for the ability to resolve mixtures of proteins with known charge differences.
(8) RIEF may be more efficient due to the documented electrodissociation of noncovalent protein:protein complexes which occurs during RIEF separations.
(9) It is concluded that the borate-glycerol system can be adequately stabilized in RIEF for isoelectric purification of certain proteins.
(10) Fibrolase, a blood clot-lysing enzyme, was isolated from the venom of the snake Agkistrodon contortrix contortrix using preparative scale isoelectric focusing in the recycling isoelectric focusing (RIEF) apparatus.
(11) Both CFE and RIEF apparently separated the components of both mixtures into individual peaks, separated by fractions which contained little or no detectable protein.