Preparation and analysis of bifunctional immunoconjugates containing monoclonal antibodies OKT3 and BABR1

Summary OKT3 and BABR1 anti-(breast tumour) antibody] were conjugated usingN-succinimidyl-3-(2-pyridyldithio)propionate (SPDP). The procedure employed mild reducing conditions for SPDP-BABR1 and short conjugation incubations at 37°C. The bifunctional immunoconjugates thus produced were isolated by HPLC gel filtration on a preparative TSK 3000 SW column. Both intact IgG and F(ab)₂ fragments have been conjugated. The ratio of SPDP to IgG for the optimal yield of dimeric OKT3-BABR1 heteroconjugates was determined to be 2 for OKT3 and 1–2 for BABR1. The OKT3-BABR1 dimers were shown to be bifunctional heteroconjugates by polyacrylamide gel electrophoresis, isoelectric focusing, radial immunodiffusion, and flow cytometry. The binding specificities of the bifunctional heteroconjugates were identical to the specificities of both parent antibodies.