Development Of An Indirect Competitive Chemiluminescence Enzyme-linked Immunoassay Method For Zearalenone

As an estrogen biological activity of the mycotoxin, zearalenone has thereproductive development toxicity, immunosuppression toxicity, hepatic toxicity,cellular toxicity and hazards tumor formation, and so on. Not only to bring hugeeconomic loss for breeding industry, but also to harm health of human and animalsseriously. Therefore, to develop a rapid detection method for zearalenone, which hasa low detection limit, high sensitivity and accuracy, is of great significance toguarantee food and feed safety. An indirect competitive enzyme-linkedimmunosorbent assay (ELISA) method and chemiluminescence enzyme-linkedimmunoassay (CLEIA) method using the monoclonal antibody were developed toanalyze zearalenone. Meanwhile, the chemiluminescent enzyme-linkedimmunosorbent kit was simple assembled. The results would be contributed to theprogress of rapid and quantitative detection methods for zearalenone.The hybridoma cell strain, which was storaged by the laboratory, secretingmonoclonal antibody against zearalenone was screened by subclone. The asciteswere prepared in vivo of mice inducing. The biological characteristics againstzearalenone monoclonal antibody were identified. The subclass of the monoclonalantibody was IgG1. The ascites were purified by the Ammonium Sulfate-OctanoicAcid combined with protein G affinity chromatography method. The ELISA titerwas1:6.4×10~4, and the affinity constant was2.238×109L/mol. The cross reactivitieswith coupling protein and homoplastic mycotoxin were less than0.01%.An indirect competitive ELISA method for detection zearalenone using themonoclonal antibody as the foundation has been established and optimized. Thelinear regression equation of standard curve was y=-23.908x+64.266(Thecorrelation coefficient R2=0.9851). The linear range was0.220ng/mL~71.042ng/mL,the sensitivity was3.951ng/mL, and the lowest detection limit was0.136ng/mL. Thespecificity, precision and the spiked corn samples recovery experiment of detection method were identified well.An indirect competitive CLEIA method for detection zearalenone using themonoclonal antibody and the indirect competitive ELISA method establishedpreliminary as the foundation has been established and optimized. The linearregression equation of standard curve was y=-19.308x+36.899(The correlationcoefficient R2=0.9692). The linear range was0.00586ng/mL~7.503ng/mL.Thesensitivity was0.210ng/mL and the lowest detection limit was0.00323ng/mL.The relevant technical parameters of simple assembled CLEIA kit were studied,the results showed that it remains to perfect, in order to improve the accuracy,precision and stability.