测定司帕沙星间接竞争酶联免疫检测方法建立及应用  

Establishment of a competitive inhibition enzyme-linked immunosorbent assay for determination of sparfloxacin and its applications

作　　者：曾华金[1] 刘兵[2] 杨冉[2] 雷利芳[2] 屈凌波[2,3] 

机构地区：[1]郑州大学药学院,郑州450001 [2]郑州大学化学系,郑州450001 [3]河南工业大学化学化工学院,郑州450001

出　　处：《中国抗生素杂志》2011年第7期530-535,共6页Chinese Journal of Antibiotics

摘　　要：目的建立快速、灵敏的司帕沙星残留间接竞争酶联免疫检测方法。方法采用混合酸酐法,将司帕沙星与牛血清白蛋白和卵清蛋白分别合成免疫抗原(sparfloxacin-BSA)和包被抗原(sparfloxacin-OVA)。通过背部多点免疫法免疫大白耳兔,制备了抗司帕沙星的特异性抗血清。结果利用所获得的特异性抗血清,建立了测定司帕沙星的间接竞争酶联免疫检测方法(ci-ELISA),其线性范围为5ng/mL～2μg/mL(R2=0.9942),且与HPLC方法具有良好的相关性(R2=0.9918)。样品牛奶、大鼠血浆和尿液中的加样回收率分别为93.7～110.3%、105.6～113.3%和90.2～98.1%%。应用所建立的方法,对司帕沙星大鼠体内药物分析及药品中的含量进行了测定。结论本研究所建立的检测司帕沙星残留的酶联免疫检测方法具有样品处理简单,灵敏度高及分析速度快等优点,非常适合于生物样品分析及药品残留的筛选工作。Objective To establish a rapid and sensitive competitive inhibition enzyme-linked immunosorbent assay （ci-ELISA） for determination of sparfloxacin （SPFX）. Methods Immunogen sparfloxacin-bovine serum albumin （SPFX-BSA） and coating antigen sparfloxacin-ovum albumin （SPFX-OVA） were prepared by a succinic anhydride method, respectively. By subcutaneous injection, the polyclonal antiserum was produced from big ear rabbits immunized with conjugates SPFX-BSA. Results As a reagent, the antiserum was also determined with a competitive inhibition Enzyme-linked Immunosorbent Assay （ci-ELISA） method and the working range of this method extended from 5ng/mL-2μg/mL （R^2=0.9942）. In a comparison of the assay results obtained by ELISA and high performance liquid chromatography （HPLC）, there is a good correlation between these two methods （R^2=0.9918）. The proposed method has been satisfactorily applied for the determination of SPFX in rat plasma and pharmaceuticals with recoveries in the range of 93.7% to 110.3% for milk sample, 105.6%-113.3% for rat plasma and 90.2%-91.8% for rat urine. Conclusion The experimental data indicated that in some extent the ci-ELISA method exhibits more advantages over other methods, such as lower detection limit, low background and no requirement of sample pretreatment, and more suitable for high throughput and real-time SPFX analysis in biosamples and pharmaceuticals.

关 键 词：司帕沙星 抗血清 ci—ELISA 

分 类 号：R917[医药卫生—药物分析学]