Studies on Aqueous Two Phase Polymer Systems Useful for Partitioning of Biological Materials
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STUDIES ON AQUEOUS TWO PHASE POLYMER SYSTEMS USEFUL FOR PARTITIONING OF BIOLOGICAL MATERIALS
DONALD E. BROOKS*+ AND STEPHAN BAMBERGER* *Department of Neurology, University of Oregon Health Sciences Center, Portland, Oregon and +Departments of Pathology and Chemistry, University of British Columbia, Vancouver, Canada
ABSTRACT The two phase systems that result when aqueous solutions of dextran and poly(ethylene glycol) (PEG) are mixed above a critical concentration of a few percent provide a useful medium for the separation of biological cell subpopulations via partition between the top, PEG-rich phase and the liquid-liquid phase boundary. Interfacial tensions of such systems have been measured by the rotating drop technique and found to range between 10-1 and 102 UN 1 m- . The tension was found to depend on the length of the tie line describing the system on a phase diagram, via a power law relationship which differed depending on the concentration of Na phosphate buffer present. The electrokinetic properties of drops of one phase suspended in the other were studied for a variety of systems. It was found that the droplet electrophoretic mobility increased monotonically with phosphate concentration and drop diameter but exhibited the opposite sign from that anticipated from phosphate partition measurements. It was possible to take advantage of these electrokinetic properties and dramatically enhance the speed of phase separation through application of relatively small electric fields. INTRODUCT ION The two phase systems which form when dextran (poly(c-l,6 glucose)) and poly(ethylene glycol) (PEG) are mixed to form aqueous solutions which are greater than a few percent in each, when appropriately buffered, have proven to be valuable as partition media for macromolecules, sub-cellular organelles and whole biological cells [1-3]. When cells or microscopic particles are introduced to such systems, the systems shaken to emulsify the phases, and allowed to re-settle, it is frequently found that the cells are distributed between the top, PEG-rich phase and the interface between the bulk phases, the dextran-rich phase being empty of cells. The interface therefore acts as a third phase with respect to the distribution of particulates in the systems. The strength of this procedure as an analytical or preparative process derives from the fact that the partition coefficient (percentage of cells found in the top phase) is determined, ideally, by cell membrane properties which are under direct genetic control and which can vary in ways relevant to cell activity and function. While for macromolecules the partition coefficient is determined purely thermodynamically, for particulates such as cells which do not diffuse to any great degree because of their size (1-50 Um diameter) the determining mechanisms are not well understood.
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Partitioning works well for relatively small cells.
An inherent
limitation appears for cells which sediment significantly during the time required for the phases to settle, however, since such
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