A thyroglobulin conjugate of dioxin (thyroglobulin-2 adipamide, 3,7,8-trichlorodibenzo-p-dioxin) (TG-TCDD) was used to immunize BALB/c mice. Hybridomas were produced by cell fusion between immune spleen cells and mouse myelomas SP2/0, P3, or NS1. In order to screen the thousands of resultant cultures for production of MoAb, a rapid, solid phase radioimmunoassay for antibody to dioxins was developed. This involved attaching bovine serum albumin coupled with trichlorodibenzo-p-dioxin (BSA-TCDD) to polystyrene plates to be used as a solid phase target antigen for reaction with MoAb. Fourteen hybridomas were identified that produced MoAb reacting with BSA-TCDD but not with BSA alone. Antibodies were tested for binding to BSA-aniline to eliminate those with limited binding specificity. Initial studies indicated that most MoAbs bound BSA-aniline as well as BSA-TCDD. More detailed analyses indicated that while most MoAbs showed some reaction with BSA-aniline, two showed preferential binding to BSA-TCDD of more than 200 fold whereas rabbit antisera demonstrated only a 5-fold discrimination. MoAb 391-1B was purified from mouse ascites fluid and after radioiodination, was tested for direct binding to BSA-TCDD or BSA-aniline. 125I MoAb showed no significant binding to BSA-aniline while demonstrating high binding to BSA-TCDD (Ka = 4.5 × 1081 mole). Experiments are in progress to use these MoAbs to develop sensitive radioimmunoassays or enzyme linked assays specific for 2,3,7,8-TCDD.
ASJC Scopus subject areas
- Environmental Engineering
- Environmental Chemistry
- Health, Toxicology and Mutagenesis