Glutaramyl-.beta.-alanyl Spacer Group for Haptenic Coupling to Proteins. Preparation of Immunogens for Antibody Production against Polychlorinated Biphenyls

By use of a glutaramyl-beta-alanyl spacer group, a hapten for the polychlorinated biphenyl, 2,2',4,4',5,5'-hexachlorobiphenyl (1), viz., 2-amino-2',4,4',5,5'-pentachlorobiphenyl (2), was successfully conjugated to carrier proteins to provide immunogens with high hapten/protein molar substitution ratios (MSR's). The procedure allows for the incorporation of beta-[3H]-alanine into the immunogen, thereby providing an accurate radiochemical method for the quantitative assessment of MSR. The use of the glutaramyl spacer group was prompted by the observation that the corresponding succinamyl group was subject to side reactions manifested by succinimide formation during the carboxyl activation step to an activated ester for subsequent coupling to proteins, thus severely compromising the coupling yields. The glutaramyl-beta-alanyl spacer group should be generally applicable for protein conjugation of any hapten with an amino functional group in the molecule.