玉米赤霉烯酮单克隆抗体和免疫酶技术研究

采用蛋白质连接技术合成玉米赤霉烯酮抗原,免疫Balb/c鼠,通过淋巴细胞杂交瘤技术建立六株分泌抗玉米赤霉烯酮的单克隆抗体杂交瘤细胞株。间接酶联免疫吸附试验测定细胞上清抗体效价为1:2084(4H8)、1:256(6H9、4H3、2H5、2C8)、1:16(3F10);腹水抗体效价为10~9(4H3、4H8)、10~8(2H5)、10~7(6H9)、10~5(3H10)。竞争间接酶联免疫吸附试验测定六株单克隆抗体对玉米赤霉烯酮的敏感度为0.3—0.8ng/ml。六株抗体与玉米亦霉烯醇的交叉反应率为1.3—9.0％。六株单克隆抗体均属IgG类。细胞体外传代培养和冻存复苏后分泌抗体稳定。纯化抗体在37℃保存12天稳定,-30℃保存90天抗体滴度不变。用该抗体建立竞争间接酶联免疫吸附试验检测掺合玉米赤霉烯酮的玉米、小麦、饲料,平均回收率分别为105％、90％、103％,平均批间变异系数为5.8％、2.8％、6.8％,批内变异系数为3.8％、12.7％、15.7％。样品中玉米赤霉烯酮掺合量与竞争间接酶联免疫吸附试验检出量有良好相关性(r≥0.9996)。

MONOCLONAL ANTIBODIES AGAINST MYCOTOXIN ZEARALENONE AND ITS APPLICATION IN IMMUNOASSAY

Six stabilized hybridoma cell lines secreting anti-zearalenone (ZEN) monoclonal antibodies (IgG) were produced by fusing the mouse cells (SP2/0) with splenocytes isolated from Balb/c mice. The Balb/c mice were previously immunized with ZEN-bovine serum albumin (BSA) conjugation which was prepared by ZEN-oxime with BSA. An indirect competitive ELISA was used to detect ZEN by anti-ZEN McAb. Cross reactivity of these antibodies with zearalenol was 1.3-9.0%. The sensitivity of CI-ELISA for ZEN was 0.3-0.8ng/ml. Average recovery rates of ZEN spiked (25-500ng/g) corn, feed and wheat were 105%, 103% and 90%, respectively. Mean coefficients of variation were 3.8%, 15.2% and 12.7% for interwell and 5.8%, 6.8% and 2.8% for interassay, respectively.