Production And Identification Of Ractopamine Artificial Antigen

Classed as phenolic amine, ractopamine (ractopamine, RAC) is a β2adrenergicreceptor agonists with effects of nutrition redistribution and growth promotion,similarly to the “lean” clenbuterol, which is used as additive to increase lean musclegrowth and protein accretion. However, the residues of RAC in animal and theserious harm to people ’s health, which may pose a potential risk for consumer health,resulting in some harmful effects on human health including headache, dizziness,chest distress, heart palpitations, and limb numbness.In China, the production and sale of ractopamine has baned while the illegal useof ractopimine is occasionally reported. Currently, more immunological analysismethods are used to detect residual RAC, for example, by enzyme-linkedimmunosorbent assay (ELISA) for screening of RAC residue, and then confirmed bymass spectrometry or chromatography. Benefit from high sensitivity, high throughput,lower cost and ease of operation, immunological detection of RAC residue is nowwidely used, compared with HPLC, mass spectrometry or biosensors methods.In this study, we prepared ractopamine artificial antigen by three differentmethods. We also prepared the artificial antigen by the method of1,4-butanediolether (method of Linker) and selected the RAC-OVA as the coating antigen. Inaddition, We repared the artificial antigen by the method of mixed acid anhydride,and the coupling ratio of RAC-BSA is8:1,17:1and23:1. Moreover, this is the firststudy to prepare RAC artificial antigen by the methord of N’N-carbonyldiimidazole,using different solvents to get two concentrations and coupling ratios of RAC-BSAartificial antigen.Physical (UV spectrum scanning identification and SDS-PAGE gelelectrophoresis) and immunological methods (indirect ELISA) were applied toidentify the anticipated coupling of artificial antigen.In this study, we obtained RAC artificial antigen by three different methods. It isthe first study to produce RAC artificial antigen by the method ofN’N-carbonyldiimidazole, with the antisera titer about1:6400. This study laid a solid foundation for the production of highly specific monoclonal anti-RAC antibodies.