抗日本血吸虫SEA特异性IgY应用于循环抗原检测的研究  

Detection of Circulating Schistosome Antigen with Anti-SEA Specific Egg Yolk Antibody （IgY）

作　　者：楼涤[1] 陆绍红[1] 陈睿[1] 童群波[1] 丁建祖[1] 卓敏敏[1] 

机构地区：[1]浙江省医学科学院寄生虫病研究所,杭州310013

出　　处：《寄生虫与医学昆虫学报》2011年第4期193-197,共5页Acta Parasitologica et Medica Entomologica Sinica

基　　金：浙江省卫生厅项目（2007A003）;浙江省科技计划项目（2009F20036）;浙江省医学重点学科群项目（XKQ-009-003）：浙江省卫生高层次创新人才培养工程项目

摘　　要：本研究应用抗日本血吸虫可溶性虫卵抗原（SEA）的鸡卵黄免疫球蛋白（IgY）建立敏感、特异的检测循环抗原的双抗体夹心酶联免疫吸附试验（ELISA）。用SEA皮下多点注射法免疫海蓝鸡，水稀释法制备IgY抗体，以辣根过氧化物酶标记纯化的IgY抗体（IgY·E）和兔抗IgG抗体（IgG—E）分别作为检测抗体，IgY抗体和兔抗IgG抗体分别作为捕捉抗体，组合建立4种双抗体夹心酶联免疫吸附试验法（ELISA）检测循环抗原，分析各种方法检测SEA的灵敏度，并平行检测急性日本血吸虫病人血清6份、慢性日本血吸虫病人血清29份、华支睾阳性者血清9份、卫氏并殖吸虫阳性者血清15份、弓形虫阳性病人血清7份、正常体检者血清118份。结果显示，以IgY为捕捉抗体或检测抗体，可检测到浓度为7．5ng／mL的SEA，灵敏度是基于IgG的夹心ELISA的4倍。4种夹心ELISA法检测急血病人血清阳性率均为100％，检测正常人血清的特异性为99．1％；与弓形虫、华支睾吸虫阳性者血清无交叉反应；-而检测慢血病人血清和并殖吸虫阳性者血清时，以IgY为捕捉抗体的2种ELISA（IgY＋IgY—E、IgY＋IgG—E）阳性率均分别为79．3％、6．7％；而IgG为捕捉抗体时，以IgY为检测抗体（IgG＋IgY—E）时的阳性率分别为75．8％、6．7％，以IgG为检测抗体（IgG＋IgG-E）时的阳性率分别为72．4％、13．3％，以IgY作为捕捉抗体的检测效果优于IgG。基于IgY抗体的双抗体夹心ELISA检测循环抗原具有良好的敏感性和特异性，可用于日本血吸虫病的免疫诊断。A sensitive and specific double-antibody sandwich enzyme-linked immunosorbent assay （ELISA） was developed to detect circulating schistosome antigen with chicken egg yolk immunoglobulin antibodies （IgY） against soluble egg antigen （SEA）. Hens were subcutaneously immunized with SEA and the crude IgY was extracted from egg yolk by water dilution method. Purified IgY antibodies and rabbit IgG antibodies against S. japonicum were labeled with horseradish peroxidase respectively （IgY-E and IgG-E）. 4 double-antibody sandwich ELISA methods were developed using IgY or IgG as capture antibody and using IgY-E or IgG-E as detecting antibody respectively. The detection limits of SEA of 4 ELISA methods were analyzed. Then human serum from patients and healthy people were detected in parallel by established 4 ELISA methods, including 6 serum samples from acute schistosomiasis patients, 29 from chronic schistosomiasis patients, 118 from healthy people, 9 from clonorchiasis patients, 15 from paragonimiasis patients, and 7 from toxoplasmiasis patients. The results showed that the minimal detectable concentration of SEA was 7.5 ng/ml by IgY based-sandwich ELISA, the sensitivity was higher than IgG based-sandwich ELISA by 4 times. All sandwich ELISA methods showed a sensitivity of 100% for acute schistosomiasis patients, a specificity of 99. 1% for healthy persons; no cross-reaction with Toxoplasmiasis and clonorchiasis; The positive rate of samples from 29 chronic schistosomiasis patients and 15 paragonimiasis patients were 79.3% , 6.7% respectively when using IgY as capture antibody, while the positive rate were 75.8% , 6. 7% and 72.4% , 13.3% when using IgG as capture antibody, IgY-E and IgG-E as detecting antibody respectively, indicating the assay involved the use of IgY was better than that of rabbit anti-schistosoma IgG. The novel double-antibody sandwich ELISA assay using anti-SEA IgY appears to be sensitive and specific for detection of circulating schistosome antigens. It is the promising assay for immunodi

关 键 词：IGY 日本血吸虫 循环抗原 双抗体夹心ELISA 

分 类 号：S858.235.9[农业科学—临床兽医学]