黄曲霉毒素M1；单克隆抗体；酶联免疫吸附试验_

抗黄曲霉毒素M1抗体制备及检测方法建立

http://www.100md.com 《中国公共卫生》 2007年第1期

黄曲霉毒素M1；单克隆抗体；酶联免疫吸附试验,,黄曲霉毒素M1；单克隆抗体；酶联免疫吸附试验,1材料与方法,2结果,3讨论,参考文献

     摘要：目的制备针对黄曲霉毒素M1的单克隆抗体并建立针对黄曲霉毒素M1的间接竞争酶联免疫吸附试验检测方法。方法利用B细胞杂交瘤技术，建立能分泌抗黄曲霉毒素M1单克隆抗体的杂交瘤细胞株，制备抗黄曲霉毒素M1单克隆抗体，建立间接竞争酶联免疫吸附试验检测方法。结果研制出1株能特异性分泌抗黄曲霉毒素M1单克隆抗体的杂交瘤细胞株，命名为2F2。该单克隆抗体的Ig亚类为IgG1，亲和常数为28×10-11mol/L。该抗体与黄曲霉毒素B1、黄曲霉毒素B2、黄曲霉毒素G1、黄曲霉毒素G2和黄曲霉毒素M2等结构类似物有微弱的交叉反应，具有较高的特异性。在此基础上建立了间接竞争酶联免疫吸附试验检测方法。该方法的最低检出浓度为007ng/ml，校正曲线的线性范围为002～2ng/ml，线性方程y=-04364x+02693(R2=09949)。方法的加标回收率为725%～1313%。结论制备了具有高特异性和亲和力的抗黄曲霉毒素M1单克隆抗体，并建立了快速、灵敏的针对黄曲霉毒素M1的酶联免疫吸附试验检测方法。

    关键词：黄曲霉毒素M1；单克隆抗体；酶联免疫吸附试验

    Development of monoclonal antibody against aflatoxin M1 and immunoassay for aflatoxin M1

    JIANG Tao,YU Qiong,LI Min,et al.

    National Institute for Nutrition and Food Safety,Chinese Center for Disease Control and Prevention(Beijing 100021,China)

    Abstract： Objective To prepare monoclonal antibody against aflatoxin M1 and develop indirect competitive enzyme-linked immunosorbent assay for detection of aflatoxin M1.Methods Hybridoma cell line excreting monoclonal antibody against aflatoxin M1 was produced using B cell hybridoma technique and develop indirect competitive enzyme-linked immunosorbent assay for detection of aflatoxin M1.Results One hybridoma cell line excreting monoclonal antibodies against aflatoxin M1 coded 2F2 was obtained by fusing murine Sp2/0 cells with spleen cells from BALB/c mice immunized with AFM1-BSA conjugate.The monoclonal antibody produced by the hybridoma cell were tested for subtype and designated as IgG1 for 2F2.The affinity of IgG in purified ascites yielded by hybridoma cell was 28×10-11 mol/L.The monoclonal antibody obtained in the present study was specific to aflatoxin M1,because of lightly cross reactions among the monoclonal antibodies against aflatoxin M1 with the analogues of aflatoxin B1,aflatoxin B2,aflatoxin G1,aflatoxin G2 and aflatoxin M2 were found.The limit of detecting concentration of aflatoxin M1 was 0.07 ng/ml.The linear range of standard curve was 002～2ng/ml and the linear equation was y=-04364x+02693(R2=0.9949).The recovery of aflatoxin M1 was between 725%～1313%.Conclusion The monoclonal antibody obtained in the study was of relatively high specificity to aflatoxin M1,and a simple fast sensitive indirect competitive enzyme-linked immunosorbent assay for the detection of aflatoxin M1 was developed. ......

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