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目的:建立快速、灵敏的恩诺沙星残留酶联免疫检测方法(ELISA)。方法:采用混合酸酐法,将恩诺沙星与牛血清白蛋白和卵清蛋白分别合成免疫抗原(Enrofloxac in-BSA)和包被抗原(Enrofloxac in-OVA)。通过背部多点免疫法免疫日本大白耳兔,制备抗恩诺沙星的特异性抗血清。结果:利用所获得的特异性抗血清,建立的恩诺沙星ELISA检测方法,其线性范围为5 ng.mL-1～2.5μg.mL-1(R2=0.9935),且与HPLC方法具有良好的相关性(R2=0.9892,n=9)。牛奶、大鼠血浆和尿液中的加样回收率分别为98.4%～105.8%,91.7%～101.2%,97.0%～110.3%。运用所建立的方法,对大鼠体内血浆及药品中恩诺沙星的含量进行了测定。结论:本研究所建立的检测恩诺沙星的ELISA方法具有样品前处理简单,灵敏度高及分析速度快等优点,适合用于恩诺沙星的体内分析及动物性食品中恩诺沙星的残留分析。
Objective: To establish a rapid and sensitive Enrofloxacin Residual Enzyme Linked Immunosorbent Assay (ELISA). Methods: Enrofloxacin-BSA and Enrofloxac in-OVA were respectively synthesized by enantioin, bovine serum albumin and albumin by mixed acid anhydride method. The immunized Japanese white-ear rabbits were immunized with multi-point immunization to prepare specific anti-enrofloxacin antiserum. Results: Enrofloxacin ELISA was established with the specific antiserum obtained. The linear range was 5 ng.mL-1 ~ 2.5 μg.mL-1 (R2 = 0.9935), which was in good agreement with the HPLC method (R2 = 0.9892, n = 9). The recoveries of milk, rat plasma and urine were 98.4% -105.8%, 91.7% -101.2% and 97.0% -110.3%, respectively. Using the established method, the contents of enrofloxacin in plasma and medicine of rats were determined. Conclusion: The enzyme-linked immunosorbent assay (ELISA) for the detection of enrofloxacin established in this study has the advantages of simple sample pretreatment, high sensitivity and fast analysis. It is suitable for in vivo analysis of enrofloxacin and enrofloxacin in animal food Residue analysis.