PRODUCTION OF HYBRIDS SECRETING BISPECIFIC ANTIBODIES RECOGNIZING CEAAND DOXORUBICIN

A monoclonal anti-CEA secreting hybridoma (11-285-14) was made hypoxan thine, aminopterin and thymidine (HAT) sensitive by back selecting it in increasing concentrations of 8-azaguanine. Eight 8-azaguanine resis tant fusion partners were selected based an growth characteristics and continued anti-CEA production. Since doxorubicin (Dox) is a hapten, i t was conjugated to the carrier proteins keyhole limpet hemocyanin (KL H) or bovine serum albumin (BSA) using 1-ethyl-3- (dimethyl-aminopropy l) carbodiimide. Dox-KLH and Dox-BSA conjugates were used to immunize mice and spleen cells from these mice were used for fusions with the H AT sensitive anti-CEA partner using standard hybridoma procedures. Enz yme linked immunosorbent assays (ELISAs) were developed to test the hy brids obtained for anti-CEA, anti-DOX, anti-BSA and bispecific monoclo nal antibody (BsMab) activity. Sixteen fusions with spleen cells from DOX-KLH immunized mice yielded 621 hybrids of which 47 showed low leve l BsMab activity by ELISA. Eight fusions with spleen cells from DOX-BS A immunized mice yielded 297 hybrids. Fifty of these hybrids showing d ual reactivity have been cloned and subcloned to yield 7 subclones wit h stable BsMab activity for CEA and doxorubicin.