Improved conditions for the production of monoclonal antibodies to carcinogen-modified DNA, for use in enzyme-linked immunosorbent assays (ELISA).
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 ring-opened aflatoxin B1-modified DNA as an example (iro-AFB1 DNA). The percentage of immunised mice which responded to iro-AFB1 DNA-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-AFB1 DNA 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-AFB1 DNA was used. Other factors found to improve the number and titre of mice responding to immunisation and the yield of hybridomas were: KLH greater than BSA as carrier protein, C57 BL/6 X BALB/c F1 greater than 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 beta-galactosidase-linked sheep F(ab')2 anti-mouse IgG as the second antibody were also tested. Present experience with AFB1 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.
D016604:Aflatoxin B1; D000348:Aflatoxins; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics* / Q000302:isolation & purification; D000936:Antigen-Antibody Complex; D002273:Carcinogens; D002417:Cattle; D002460:Cell Line; D004247:DNA / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D006825:Hybridomas / Q000276:immunology; D008214:Lymphocytes / Q000276:immunology; D051379:Mice; D010954:Plasmacytoma / Q000276:immunology; D013950:Thymus Gland
D000348:Aflatoxins; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D002273:Carcinogens; D004247:DNA; D016604:Aflatoxin B1