title: Comparison of methods for the generation of immunoreactive fragments of a monoclonal antibody (B72.3) reactive with human carcinomas.
abstract: Immunoglobulin fragments, whether of polyclonal or monoclonal antibodies, offer a number of advantages over the intact immunoglobulin. The generation of immunoreactive fragments from monoclonal antibodies (MAb) is not always a straightforward task. Both pepsin and papain can be used to digest MAb to a bivalent molecule with a Mr of 100,000. However, pepsin pepsin digestions does not always result in immunoreactive fragments and a stable consistent product by papain digestion is often difficult to obtain. MAb B72.3 is an example of both situations. MAb B72.3 reacts with a glycoprotein (TAG-72) with a molecular weight greater than 10(6). MAb B72.3 has been shown to exhibit a high degree of selective reactivity with colon, breast and ovarian carcinomas and has been used for radioimmunodiagnosis in model systems and in clinical trials. A third enzyme, bromelain, in the same family of sulfhydryl proteases as papain, has been used to generate a fragment of MAb B72.3, with a Mr of approximately 100,000. The bromelain-generated fragment of MAb B72.3 retained 100% immunoreactivity as measured in competitive solid-phase radioimmunoassays and could be generated with consistent results from one preparation to another. Both the bromelain- and papain-generated fragments were radiolabelled with 125I without significant loss of the MAb's reactivity to tumor extracts. Differences were observed between the bromelain- and papain-generated fragment when compared in vivo. Fragmentation of MAb B72.3 with bromelain has yielded a superior bivalent fragment for radioimmunolocalization.
authors.affiliation: Laboratory of Tumor Immunology and Biology, National Cancer Institute, Bethesda, MD 20892.
authors.forename: D E
authors.initials: DE
authors.lastname: Milenic
authors.forename: J M
authors.initials: JM
authors.lastname: Esteban
authors.forename: D
authors.initials: D
authors.lastname: Colcher
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000493:pharmacokinetics / Q000627:therapeutic use; D000912:Antibodies, Neoplasm / Q000276:immunology* / Q000493:pharmacokinetics; D001963:Bromelains; D002460:Cell Line; D007128:Immunoglobulin Fragments / Q000276:immunology* / Q000302:isolation & purification; D007167:Immunotherapy; D051379:Mice; D008819:Mice, Nude; D008970:Molecular Weight; D009374:Neoplasms, Experimental / Q000000981:diagnostic imaging / Q000628:therapy; D010206:Papain; D010434:Pepsin A; D011877:Radionuclide Imaging
chemical_list: D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; D007128:Immunoglobulin Fragments; D001963:Bromelains; D010206:Papain; D010434:Pepsin A