Improved conditions for the production of monoclonal antibodies to carcinogen-modified DNA, for use in enzyme-linked immunosorbent assays (ELISA)

Abstract

The methodology for the production of monoclonal antibodies to chemical carcinogen-modified DNA has been improved to provide high yields of hybridomas, using guanine-imidazole rine-opened aflatoxin B 1-modified DNA as an example (iro-AFB 1DNA). The percentage of immunised mice which responded to iro-AFB 1DNA-protein immunisation and the number of specific hybridomas produced was dependent on the level of modification of DNA. One in three BALB/c mice had detectable (but low) antibody titre when 0.3% modified iro-AFB 1DNA was used and this yielded 2 specific hybridomas, whereas all mice responded at reasonable titres and 6 specific hybridomas were obtained when 3% modified iro-AFB 1DNA was used. Other factors found to improve the number and titre of mice responding to immunisation and the yield of hybridomas were: KLH > BSA as carrier protein, C57 BL/6 × BALB/c F1 > BALB/c mice for antibody production, fusion success and ascites growth. The conditions limiting the sensitivity and reproducibility of an enzyme-linked immunosorbent assay (ELISA) using these monoclonal antibodies with β-galactosidase-linked sheep F(ab′) 2 anti-mouse IgG as the second antibody were also tested. Present experience with AFB 1 and other carcinogens indicates that these methods should be applicable to the production of monoclonal antibodies to DNA modified by a wide variety of chemical carcinogens.