Em18重组抗原特异性抗体的制备及对循环抗原检测的初探  

Preparation of Specific Antibodies Against Recombinant Em18 Antigen and Detection of the Circulating Antigen

作　　者：江莉[1] 李雄[2] 李浩[1] 牛新玲[2] 冯正[1] 

机构地区：[1]中国疾病预防控制中心寄生虫病预防控制所世界卫生组织疟疾,血吸虫病和丝虫病合作中心 [2]新疆维吾尔自治区疾病预防控制中心,乌鲁木齐830002

出　　处：《中国寄生虫学与寄生虫病杂志》2006年第5期329-332,共4页Chinese Journal of Parasitology and Parasitic Diseases

摘　　要：目的用Em18重组抗原ReEm18制备特异性抗体,建立双抗体夹心ELISA,检测患者血清中的循环抗原。方法ReEm18经亲和纯化,免疫BALB/c小鼠和新西兰兔,分别制备单克隆抗体(单抗)和多克隆抗体(多抗)。用获得的单抗和多抗建立双抗体夹心ELISA,检测患者血清中Em18循环抗原。结果免疫兔血清抗ReEm18抗体效价达1∶204800以上。经ReEm18和多房棘球蚴病(AE)患者血清及健康人血清阻断ELISA试验双重筛选融合细胞,共获得14株抑制率>50%的特异性阳性细胞克隆。将特异性单抗和多抗自由组合进行双抗体夹心ELISA,结果以9号单抗与多抗的配对检测抗原为最优,检测ReEm18的灵敏度达3ng/ml。用建立的双夹心ELISA方法检测血清,11份AE血清中6份为阳性。结论获得针对ReEm18的特异性单抗和多抗,建立了敏感的双抗体夹心ELISA方法,血清学初步检测结果表明AE血清中存在可检测的Em18循环抗原。Objective To prepare specific antibodies against the recombinant Em18 antigen （ReEm18）, estab-lish a sandwich ELISA and detect the Em18 circulating antigen in patients＇ sera. Methods Rabbits and BALB/c mice were immunized with the ReEm18 antigen, which was purified by ＇affinity method for preparation of the specific polyclonal and monoclonal antibodies. A sandwich ELISA was established by the specific antibodies. Results By immunizing with the ReEm18 antigen, high antibody level was reached with a serum dilution of 1：204 800 and above in the immunized rabbits. After double selection by ELISA using the ReEm18 antigen and block ELISA using both AE-positive and negative control sera, 14 positive cell clones were obtained with an inhibition rate of more than 50%. Those monoand poly-clonal antibodies were matched freely in sandwich ELISA tests for detecting the ReEm18 antigen. A combination of monoclonal antibody No.9 and polyclonal antibody showed the best result. The sensitivity to detect ReEm18 antigen was at 3ng/ml. Six of 11 AE sera were positive when tested with the sandwich ELISA system. Conclusion Highly specific polyclonal and monoclonal antibodies have been prepared, and a sensitive sandwich ELISA established. Preliminary result is suggested that a detectable level of Em18 circulating antigen is present in AE patients＇ sera.

关 键 词：多房棘球蚴病 Em18重组抗原 抗体 循环抗原 夹心ELISA 

分 类 号：R532.32[医药卫生—内科学] R392.11[医药卫生—临床医学]