THE CREATION OF A DOUBLE HYBRID CELL LINE SECRETING A BIFUNCTIONAL ANTIBODY MOLECULE WITH ONE BINDING SITE RECOGNIZING A MELANOMA ASSOCIATED ANTIGEN AND THE OTHER PSEUDOMONAS EXOTOXIN A

Abstract

Hybrid hybridomas (quadromas) which produced antibody of mixed parentage were isolated through the fusion of two biochemically deficient hybridoma lines. One of the parent hybridomas secreted an IgG 1 monoclonal antibody which reacted specifically to Pseudomonas exotoxin A while the other parent secreted an IgG 2b monoclonal antibody reactive to a 250 Kd melanoma associated proteoglycan. True quadromas were isolated on the basis of their ability to secrete antibody binding to both Pseudomonas exotoxin A and the proteoglycan melanoma associated antigen with the presence of both parental isotypes clearly demonstrable in the resulting immune complexes. Antibody species were separated from each other using fast protein liquid chromatography over an hydroxylapatite column. SDS-polyacrylamide gel electrophoresis demonstrated that the hybrid antibody molecules contained the heavy and light chains of both anti PE and anti proteoglycan origin. The quadroma antibody was further shown to increase the toxicity of Pseudomonas exotoxin A on melanoma cells in vitro.