Study On Enzyme Linked Immunosorbent Assay For The Chloramphenicol Based On Monoclonal Antibody

In this thesis a kind of anti-chloramphenicol monoclonal antibody and two kinds of anti-ovomucoid monoclonal antibodies were prepared. And an ELISA for detection of chloramphenicol residues in seafood was established.The chloramphenicol immunogen was made by conjugated chloramphenicol succinate activated ester to keyhole limpet hemocyanin, and was applied to immunize mice. After a fusion of the immunized mice spleen cells and SP2/0myeloma cells, cells screened and subcloned, a strain of cells which can produce anti-chloramphenicol monoclonal antibody was obtained. Ascites was produced by mice vivo culcure and purified to obtain antibody. The subtype of the antibody was identified by antibody subtype identification kit:the heavy chain was IgG1, and the light chain was Kappa. A competitive ELISA using above-mentioned antibody were developed for the detection of chloramphenicol. The competitive ELISA was performed as follows:0.02μg/well monoclonal antibody4℃coated over night,0.5%milk37℃blocked1h,0.05mol/L, pH5.7PBS diluted sample,the IC50of this kind of method was0.54±0.04ng/mL, IC15was0.10±0.03ng/mL. Cross-reactivity of this method to Chloramphenicol sodium succinate, Thiamphenicol, Florfenicol, Tetracycline and Streptomycin was78%,0.09%,0.25%,0.05%and<0.002%. Accroding to the cross-reactivity experiment, the competitive ELISA was specific to Chloramphenicol.Cod and shrimp were chosen to spike and recovery. The recovery rate were between62.2%-120%. and the coefficient of variation were between0.94%-13.52%.