呋喃它酮代谢物人工抗原的合成及抗体的制备  

Synthesis of artificial antigens and preparation for specific antisera against 3-amino-2-oxazolidinone,a metabolite of furaltadone

作　　者：束建花[1] 郭红艳[1] 王立琦[1] 高艳[1] 曾振灵[1] 

机构地区：[1]华南农业大学兽医学院,广东广州510642

出　　处：《华南农业大学学报》2014年第2期18-23,共6页Journal of South China Agricultural University

基　　金：农业部兽药质量安全监管项目(农财发[2012]36号)

摘　　要：【目的】建立高灵敏度的呋喃它酮酶联免疫检测方法.【方法】以戊二醛法将呋喃它酮代谢物5-甲基吗啉-3-氨基-2-唑烷基酮(AMOZ)与牛血清白蛋白(BSA)和卵清白蛋白(OVA)偶联,再由NaBH4还原西佛碱得到结构稳定的免疫原AMOZ-BSA和AMOZ-OVA;用对醛基苯甲酸(CPA)对AMOZ进行衍生化,得到衍生物CPAMOZ,由质谱鉴定,经N-羟基琥珀酰亚胺活化酯法将CPAMOZ与BSA和OVA偶联得到免疫原CPAMOZ-BSA和包被原CPAMOZ-OVA.经紫外扫描鉴定表明人工抗原合成成功.免疫Balb/c小鼠,ic-ELISA方法测定鼠血清多克隆抗体对AMOZ和NPAMOZ的特异性.【结果和结论】结果显示CPAMOZ-BSA免疫制备的抗体对NPAMOZ有良好的特异性.抗体效价为1∶32×104,对NPAMOZ的半抑制质量浓度为4.53 ng·mL-1;抗血清与CPAMOZ、AMOZ及呋喃它酮的交叉反应率为78.6%、1.5%及4.6%;与其他3种硝基呋喃类原药及其代谢物对硝基苯甲醛、对醛基苯甲酸等结构相似物的交叉反应均小于0.01%.[Objective]To establish a high sensitivity enzyme-linked immunosorbent assay for the detection of furaltadone .[Method]Furaltadone metabolite （ 3-amino-5-morpholinomethyl-2-oxazolidinone , AMOZ ） was coupled to carrier proteins （ bovine serum albumin or ovalbumin ） and reduced with NaBH 4 for the pro-duction of immunogen AMOZ-BSA and coating antigen AMOZ-OVA through glutaraldehyde method .With 4-carboxybenzaldehyde for derivatization reagent , AMOZ was made into CPAMOZ , which was successful-ly conjugated to carrier proteins according to the active ester method to form CPAMOZ-BSA, CPAMOZ-OVA.UV scanning showed that antigens were successfully linked to carrier proteins .After immunizing animal （ Balb/c mice） , polyclonal antibody against NPAMOZ was produced .[Result and conclusion]Antibody was diluted at 1∶32 ×10^4 and the IC50 value was 4.53 ng· mL-1 .The cross-reactivity （ CR） of the antibody with CPAMOZ, AMOZ and furaltadone was 78.6%, 1.5%and 4.6%respectively.There is almost no CR with other three nitrofurans and their metabolites , which indicates a high selectivity of the antibody .

关 键 词：呋喃它酮代谢物 半抗原 多克隆抗体 酶联免疫测定法 

分 类 号：S511[农业科学—作物学] S502