The production of a monoclonal antibody to 18-hydroxycortisol and other 18-hydroxylated steroids

Four strains of mice were immunized with 6 different conjugates of 3-O (carboxymethyl-oximino)-18-hydroxycortisol to bovine serum albumin (3 preparations), turkey serum albumin, porcine thyroglobulin, and keyhole limpet hemocyanin. Spleens from 7 of 48 mice immunized were fused with Fox/NY and/or HL-1 Friendly myeloma cell lines, yielding many positive clones for antibody formation. Short cross-reactivities were done in 293 culture supernatants and were found to have low cross-reactivity (less than 0.001%) to cortisol, but very high cross-reactivity to 18-hydroxy-11-deoxycortisol (70 to 140%). One clone showed over 100% cross-reactivity with all the 18-hydroxylated steroids studied. The major problem encountered in the generation of monoclonal antibodies was the low antibody response in the vast majority of mice injected. Half the mice developed no measurable titer, and the clones evaluated from those that did produce antibodies cross-reacted with other 18-hydroxylated steroids. Nevertheless, the antibody developed could serve in radioimmunoassay for 18-hydroxy-11-deoxycortisol separated chromatographically from other cross-reacting steroids. This is important as no synthetic 18-hydroxy-11-deoxycortisol is available.