Liquid-liquid partition chromatography as a method to examine surface properties of antibodies and antigen-antibody...

We demonstrate liquid-liquid partition chromatography in aqueous two-phase systems (LLPC) as a simple method for examining the surface properties of immunoglobulins and antigen-antibody complexes in solution. LLPC separates molecules with respect to the properties of the exposed surfaces. As an example, the method may be used to detect changes in the conformation of IgG following chemical modification like acylation or iodination. We have studied the partitioning of antibodies and antigen-antibody complexes, modelled by rabbit antibodies against three human serum proteins, in aqueous polyethylene glycol/dextran two-phase systems at pH 7. Analysis of both polyclonal and monoclonal antibodies against various antigens suggested that the partition properties of immunoglobulins are related mainly to their antigen specificity and not to subclass-specific structures. Furthermore, experiments indicated that changes in the surface properties of antigen and/or antibody following complexation may be detected. Thus, LLPC may prove to be a new way of studying the relation between antibody structure and function in solution.