Monoclonal Antibodies Against Digoxin:The Development of Specific Antibody Secreting Hybridoma Cell Lines and Antibody Characterization / 中国免疫学杂志

ABSTRACT

In order to help the clinical use and pharmacological study of cardiac glycosides, we developed eight hybridoma cell lines which could secrete specific monoclonal ant- ibodies against digoxin.Digoxin was coulped to ovalbumin(Dig-OVA)or bovine serum albumin(Dig-BSA)by the periodate oxidation method.BALB/c mice were immunized with the mixture of Dig-OVA and complete Freund's adjuvant i.p.fourteen days bef- ore fusion.From the five to two days before fusion,the mice were boosted i.p.and i.v.daily.Spleen cells from the immunized mouse were fused with SP2/0 mouse myelo- ma line.Screened by indirect ELISA using Dig-BSA as the immobilized antigen and subcloned three or four times,eight hybridoma cell lines secreting specific monoclonal antibodies against digoxin were established.Those hybridomas could also produce mo- use antibody-enriched ascites in vivo.In contrast to the immunized animale sera,all the eight monoclonal antibodies had no reaction to the carrier proteins.Results show- ed those monoclonal antibodies with different immunoglobulin H chain isotypes,five IgGl,two IgG2b and one IgM,had variant affinity to digoxin.Fine specificity determ- ination showed the seven IgG antibodies had different binding styles to ten different steroid structures,which suggested their affinity to different portion on the haptens tested.Experiment with rabbit red blood cells showed one of the high affinity mono- clonal antibodies(FD8,IgGl)could antagonize the inhibition of Na~+-K~+ ATPase activity by digoxin in vitro and in vivo.Antibody FD8 could further reverse lethal digoxin toxicity completely in Guinea pigs.It is suggested that those monoclonal antibodies could be used to develop highly sensitive immunological detection for mo- nitoring blood digitalis concentration,to study the pharmacological mechanism of ca- rdiac glycosides,as well as to rescue severe digitalis toxicity.