Research And Development Of An ELISA Rapid Test Kit For Clenbuterol

Clenbutero(lCL)is aβ-agonist registered as a veterinary bronchospasmolytic drug in some countries. It was well known that CL can be used as a growth-promoting and repartitioning agent in several livestock species at comparatively high doses, as it can improve lean meat deposition and production efficiency. However, there have been several reported incidences resulted from consumption of CL-tained edible tissues of bovine and swine. Its use as a feed additive has been banned within most countries. It is necessary to develop a sensitive and applicable assay for effective surveillance of the illicit use of CL.At present, most countries use ELISA as a rapid screen method for CL. Testing organizations in China almost all use the import kits for the testing wrok. The import kits are high in price and poor at the after sale service. The domestic kits of CL have the price advantage and can give good after sale service, but the quality problems such as poor stability, high false positive/negative rates etc are so ubiquity that the home-made kits still cannot take place of the import kits. This reseach aims at developing a self-made kit for CL which is of good stability, low false positive/negative rates and can compare with the import kits.The development of this kit bases on the principle of indirect competitive ELISA, by condition optimization,final reaction parameters are achieved: the concentration of coating antigen is 0.2μg/mL, that of antibody is 0.059μg/mL, dilution rate of IgG-HRP is 1:1000, whole reaction time is 90min. Linear range of the standard curve is 0-8ng/mL, IC50 with an average of 1.0ng/mL, LOD can reach to 0.1ng/mL on avenage, which is the same as the import kits. The self-made kit shows high sensitivity with CL and almost no cross-activity with Salbutamol, Chioramphenicol, Ractopamine and otherβ-adrenergic receptor agonists. The standard variation coefficients of intra-assay and inter-assay are 1.28-13.19% and 5-15%, respectively. Compared with the import kits(ED kits), the extraction recoveries of self-made kits are ranged from 80% to 120%, which is higher than the import kits(50%-80%); the self-made kits can also keep stable and provide reproductive result for more than 1 year, which is one-up in China and the same as the import kits. In all, on the aspects of extraction recoveries, RE of standards and veracity, the self-made kit act the same or even better than ED kits, which means the self-made kit can possiblily replace the import kits.