Preparation Of Monoclonal Antibodies Against Clenbuterol Hydrochloride And The Establishment Of An Indirect ELISA Method For Detection

Clenbuterol hydrochloride (CL) is aβ-adrenergic stimulant. It can redistribute the nutrients, promote protein synthesis and increase the lean of body, improve the feed conversion ratio. It is often illegally used as a feed additive in livestock production, but the residue is the potential human health hazards, although the use of CL is prohibited in animal production, the cases of illegal adding even causing the consumer poisoned are common be seen. In this study, the monoclonal antibodies were developed for the establishment of competitive enzyme-linked immunosorbent assay (ciELISA) for detecting CL.The diazo method was used for coupling CL with carrier protein OVA, BSA. CL-OVA was used for immunizing BALB/c mice and CL-BSA was used for detection. 4 different monoclonal antibodies were obtained. Experiments showed that supernatant of 1A3 cells were less affected by solution medium than the other four. Specificity experiment shows that 1A3 antibody has a well specificity, it reacts only with CL but the carrier protein or the hapten of ractopamine, IC50 of the 1A3 antibody against salhutamol was greater than 1000 ng/ml. Usage of 1A3 in the competitive ELISA can effectively eliminate the false-positive and false-negative ratio. Monoclonal antibody ` 1A3 were purified by Protein-A Sepharose affinity chromatography method, and IC50 of the indirect competitive ELISA was 12.0ng/ml, linear detection range of which was 0.5-100ng/ml.Monoclonal antibody 1A3 were conjugated with horseradish peroxidase (HRP), established indirect competitive ELISA was very sensitive and specific, IC50 was 3.5ng/ml, detection limit against CL was as low as 0.2ng/ml, and linear detection range was 0.2ng/ml-16ng/ml.