氟喹诺酮药物格林沙星抗体制备及其icELISA方法建立  

Development of Antibody and Indirect Competitive Enzyme-Linked Immunosorbent Assay for the Detection of Fluoroquinolone Drug Garenoxacin

作　　者：吕书为 雷红涛 唐秋实 周丽君[2] 王睿娇 卢蓝蓝 王弘 沈玉栋 孙远明 

机构地区：[1]农业部农产品贮藏质量安全风险评估实验室(广州),广东省食品质量安全重点实验室,广州五山510642 [2]中国人民解放军海军总医院,北京100037

出　　处：《中国食品学报》2014年第6期187-193,共7页Journal of Chinese Institute Of Food Science and Technology

基　　金：农业部行业公益(农业)项目资助(20100300808);国家自然科学基金(30700663,20877029);高等学校博士点基金优先发展领域项目(20114404130002);广州市科技计划项目(12C12101663)

摘　　要：目的：制备格林沙星多克隆抗体并建立间接竞争酶联免疫吸附分析方法（icELISA）。方法：采用碳二亚胺法将格林沙星偶联于牛血清白蛋白（BSA）作为免疫原,免疫新西兰大白兔获多克隆抗体;采用活泼脂法分别将格林沙星、克林沙星、加替沙星、培氟沙星和沙拉沙星偶联于卵清白蛋白（OVA）,制备5种包被抗原并做性能比较;建立icELISA方法并对其灵敏度和特异性进行评价。结果：成功制备了免疫原和包被抗原及抗体,抗体效价最高达64 000倍。以克林沙星-OVA作异源包被,建立的icELISA方法灵敏度最高,半抑制药物浓度（IC50）为9.14 ng/mL,检测限（IC10）为0.9 ng/mL,检测范围（IC20~IC80）为2.3~36.4 ng/mL,与其他喹诺酮类药物无明显交叉反应。结论：首次制备出格林沙星抗体并建立间接竞争酶联免疫吸附检测方法,该法灵敏度高、特异性强,为分析生物样品中格林沙星含量的测定提供了1种新的方法。Objective： To develop an indirect competitive enzyme-linked immunosorbent assay （icELISA） for detection of garenoxacin, anti-garenoxacin antibody and coating antigen was prepared. Methods： The immunogen was prepared with water soluble carbodiimide （EDC） method, and 5 different coating antigens with different structure were synthesized with active ester method. New Zealand white rabbits were immunized to obtain a polyclonal antibody against garenoxacin. Resuits： The heterogenous coating antigen clinafloacin-OVA was found to be most sensitive, and the antibody exhibited titer at 64 000 times. The icELISA showed highly specificity and sensitivity with the 50% inhibition concentration （IC50） of 9.14 ng/mL and the limit of detection （IC10） of 0.85 ng/mL, the dynamic detectable concentration ranged from 2.3 to 36.4 ng/mL. Conclusion： The antibody to garenoxacin and icELISA was developed for the first time with high sensitivity and specificity. This study provided a novel bioanalytical method for the detection of garenoxacin in the biological sample.

关 键 词：格林沙星 多克隆抗体 酶联免疫吸附分析方法 异源包被 

分 类 号：TQ465[化学工程—制药化工]