抗伏马菌素B_1单克隆抗体的制备及试剂盒的研制

Development of monoclonal antibody against Fumonisin B_1 and ELISA-kit of quantitative analysis for Fumonisin B_1

目的为了能够快速检测粮谷类食品中伏马菌素B1的污染水平,研制具有我国自主知识产权的针对伏马菌素B1快速检测试剂盒.方法制备了能分泌抗伏马菌素B1单克隆抗体的杂交瘤细胞株,建立了敏感、特异、快速的酶联免疫吸附试验检测方法,最终形成具有中国自主知识产权的伏马菌素B1快速检测试剂盒.结果该试剂盒所用单克隆抗体的Ig亚类为IgG1,亲和常数为8.3×10^-8mol/L.该抗体与其他结构类似物无明显交叉反应,具有较高的特异性.试剂盒对伏马菌素B1最低检出限5ng/ml,标准曲线的线性范围（50～500ng/m1）,回归方程Y=-0.582X＋1.793（r=0.99,P＜0.05）;对玉米作50ng/ml,200ng/ml和500ng/ml 3个加标浓度的回收率在71.89%～112.95%之间;试剂盒在常温状态下有效期至少10个月;实验室内的变异系数和实验室间的变异系数均小于20%.

Objective To detect the contamination level of fumonisin B1 in grain, and to develop rapid detection kit that possess patent of China. Methods Hybridoma cell llne excreting monoclonal antibody against fumonisin B1 was produced using B cell hybridoma technique and develop a rapid, sensitive, quantitative ELISA-kit for detection fumonisin B1. Results The monoclonal antibody used in the ELISA-kit were tested for subtype as IgG1 and its affinity constant was 8.3 × 10^-8 mol/L. The monoclonal antibody obtained in the present study was hizhlv snecific to fumonisin B1. because no cross reactions between the monoclonal antibodies against fumonisin B1 with the analogues of fumonisin B1 were found. The limited concentration of the ELISA-kit was 5ng/ml, linear range was 50 - 500ng/ml, the linear equation was Y= - 0. 582X ＋ 1.793（ r = 0.99, P 〈 0.05）. The recovery rate of maize on the level of 50ng/ml, 200ng/ml and 500ng/ml was among 71.89 %-112.95 %. The kit can be stored at normal tempertature in ten months at least. The coefficient of variant winthinlaboratory and between-laboratory was less than 20%.