SOLUTION-PHASE BINDING OF MONOCLONAL-ANTIBODIES TO BEE VENOM PHOSPHOLIPASE A(2)

The binding of monoclonal anti-bee venom phospholipase A(2) antibodies to their antigen was monitored by size-exclusion high performance liq uid chromatography. As judged by this panel of six antibodies, honeybe e venom phospholipase A(2) contains five binding sites, three of which are completely independent epitopes. The study revealed that this PLA (2) can accommodate three different antibodies simultaneously. The res ults demonstrate the utility of size-exclusion high performance liquid chromatography in epitope analyses, such as its ability to compare th e relative expansiveness and conformational state of the epitopes and to enumerate the antibodies that the antigen can accommodate simultane ously. The data provide compelling evidence that one of the monoclonal antibodies, M5 (which activates the enzyme), recognizes a different c onformation of phospholipase A(2) than do the other antibodies. The re sults also demonstrate that different pairs of monoclonal antibodies d iffer in their predilection to form high molecular weight complexes wi th the antigen.