Development Of Monoclonal Antibody-Based Enzyme-Linked Immunosorbent Assay Test Kit For Clenbuterol

[Objective] To develope a monoclonal antibody-Based indirect enzyme-linked immunosorbent assay (ELISA) test kit involving the synthesis of immuno-antigen of Clenbuterol (CL) and the development of monoclonal antibody (McAb) specific to CL to analyze CL in foods.[Method] The generation of immuno-antigen of CL and monoclonal antibody against CL:The conjugation of CL and protein M (the immuno-antigen of CL) was synthesized according to the method of diazotization. Balb/c mice were immunized by intrasplenic injection of CL immuno-antigen. Hybridoma cell lines producing monoclonal antibody specific to CL were established by fusion between SP2/0 myeloma cells and spleen cells from Balb/c mice immunized with conjugated antigen. The monoclonal antibody against CL was produced by immunizing animals with screened hybridoma cell lines. Its specificity was tested by the cross reactivity with CL structural analogs, such as Salbuterol, Adrenalin.The eastablisment of an indirect competitive enzyme-linked immunosorbent assay method for Clenbuterol and the development of its test kit:The coating antigen was synthesized by using the Clenbuterol linking to Ovalbumin by diazotization method, called CL-OVA. Clenbuterol was used as a kind of haptens which was competive. Both of them react with quantitive monoclonal antibody against CL. The indirect competitive enzyme-linked immunosorbent assay method for Clenbuterol and the development of its test kit were established by optimizing parameters such as the concentration of coating antigen and CL-McAb and anti-mouse IgG.. The feasibility of the kit was tested by the recovery of CL spiked in pig urine and by comparing test with the abroad Clenbuterol test kit.[Result] Three hybridoma cell lines, CL-2F12,CL-3C3,CL-4F3,producing...