作　　者： ; ; ; ; ;

机构地区： 暨南大学

出　　处： 《食品工业科技》 2013年第13期161-165,共5页

摘　　要： 目的:制备5-甲基吗啉-3-氨基-2-恶唑烷酮(AMOZ)单克隆抗体,并建立AMOZ胶体金试纸条检测方法。方法:AMOZ与对醛基苯甲酸反应生成半抗原CP-AMOZ,将CP-AMOZ与BSA、OVA偶联制备完全抗原。免疫小鼠后制备腹水型抗体,ELISA检测抗体效价与其对CP-AMOZ半抑制浓度(IC50)和最低检测限(LOD),并检测抗体的特异性。胶体金与单抗偶联,同时在NC膜上包被完全抗原、兔抗鼠多抗分别作为检测线和质控线,制备免疫层析试纸条,测定试纸条的灵敏度与特异性。结果:成功制备CP-AMOZ-BSA、CP-AMOZ-OVA完全抗原及抗CP-AMOZ的单抗,抗体效价为1∶1.6×105,对CP-AMOZ IC50为0.86μg/L,LOD0.07μg/L,与其他硝基呋喃类药物代谢物及其衍生物均无交叉反应。制备的胶体金试纸条灵敏度5μg/L。结论:成功合成了CP-AMOZ的完全抗原,免疫小鼠后制备了特异性强的单抗,并以此为基础研制出敏感、特异的胶体金快速检测试纸条。 Object = The monoclonal antibody against .5- morpholino- 3- amino-2- oxazolidone （AMOZ） was prepared, and the corresponding colloidal gold immunochromatography method was established. Method= The hapten CP-AMOZ was synthesized by the reaction between AMQZ and 4-CBA and conjugated with BSA and OVA using activated ester method.Ascitic type antibody was prepared by hybridom cell lines which secret monoclonal antibody against CP-AMQZ by mean of immunizing BALB/c mice, cell fusion, ELISA screening and subcloning. Data including the titer of antibody,its ICs0 against CP-AMOZ and LOD were detected by ELISA analysis and determine its specificity. Competitive colloidal gold immunochromatographic dipstick was produced on this base, gold nanoparticles were coupled with monoclonal antibodies, CP-AMOZ-BSA and rabbit-anti- mouse polyclonal antibody were separately coated on nitrocellulose membrane as test line and control line to detect the limit of detection and specificity.Result：The titer of the mAbs was 1：1.6 × 105,the IC50 value and LOD to CP-AMOZ were O.86μg/L and O.07μg/L respectively. The mAbs had little cross reactivity with other nitrofurans metabolites and derivates,showing high specificity to CP-AMOZ, and LOD of colloidal gold immunochromatographic dipstick was 5μg/L.Conelusion：The antigen of CP-AMOZ was successfully synthetized and the monoclonal antibody specific to CP-AMOZ was prepared,which help prepare the colloidal gold rapid detection dipstick of high sensitivity and specificity.

关 键 词： 单克隆抗体 胶体金 免疫层析

领　　域： [生物学]