【摘要】 目的制备石房蛤毒素(STX)的单克隆抗体,利用时间分辨荧光免疫分析技术建立石房蛤毒素超微量的检测方法。方法利用STX-KLH偶联抗原免疫小鼠的脾细胞与骨髓瘤细胞Sp2/0融合,筛选后获得稳定分泌抗STX抗体的杂交瘤细胞株,腹水诱生法制备大量单抗,采用间接竞争法建立STX-TRFIA时间分辨荧光免疫检测试剂盒,对试剂盒各项性能进行评价。结果该试剂盒最低检出浓度为1.0 ng/ml,校准曲线线性范围为2.0~512 ng/ml,线性方程Y=-0.455 7X+22.872(|r|=0.998 3),与腹泻性贝类毒素(DSP)、神经毒性贝类毒素(NSP)、失忆性贝类毒素(ASP)均无交叉反应,平均回收率为94.66%,与进口试剂盒同时检测广州市水产品中贝类毒素的含量,其相关系数为0.9764,P<0.001。结论试剂盒各项指标均接近进口试剂盒水平,达到检验要求,可满足实际工作需要。更多还原

【Abstract】 Objective To make monoclonal antibodies against Saxitoxin(STX)and develop a rapid, sensitive and quantitative ELISA kit for STX based on time-resolved fluoroimmunoassay technology. Methods Spleen cells from 6-8 weeks old female BALB / c mice immued with FB1-KLH were fused with the Sp2 / 0 myeloma cells. Hybridoma cell linesstably secreting Mc Ab against FB1 were selected.Ascites in mice was produced by intraperitoneal injection and antibodies were purified with protein A affinity chromatography. The STX-TRFIA detection kit was established based on indirect competitive ELISA method and evaluated. Results The limited concentration of detection was 2 ng / ml, linear range was 2-300 ng /ml; the linear equation was Y=-0.455 7X+22.872(r=0.998 3). There was no cross reaction with ASP, NSP, and DSP.The mean recovery rate was 94.66%. Compared with another kit from USA, the correlation coefficient of results from shellfish toxin levels detection in aquatic products in Guangzhou city with the kit was 0.957 4,P<0.001. Conclusion The ELISA-kit detectition ability meet the needs of practical work.更多还原