抗日本血吸虫噬菌体抗体库的构建和初步应用

Construction and Application of Phage Display Library Against Schistosoma japonicum

目的探索大规模制备抗日本血吸虫单克隆抗体的新途径。方法以日本血吸虫感染小鼠的脾细胞为源,构建抗日本血吸虫噬菌体单链抗体库,并对该抗体库进行了富集、筛选和鉴定。同时,用从抗体库制备的可溶性单链抗体对病人血清进行了检测。结果构建的噬菌体抗体库库容为1.07×106cfu。筛选获得2个特异性阳性克隆。这两株克隆表达的单链抗体用于血吸虫病人血清检测的结果为:(1)以其中一株单链抗体检测急性病人的敏感度为60%,检测慢性病人的敏感度为36.7%,特异度为90%。(2)将两株单链抗体混合后检测急性病人的敏感度为75%,检测慢性病人的敏感度为56.7%,特异度为85%。结论首次成功地构建了抗日本血吸虫噬菌体单链抗体库,并从中筛选制备了能有效检测血吸虫病患者血样中的循环抗原的单链抗体。

Objective To find a new way to produce S.japonicum (Sj) monoclonal antibodies in large scale. Methods A phage displayed antibody library against Sj was constructed with Sj infected murine spleen. To get the single chain variable fragment antibody (scFv) against Sj, the library was screened with metabolic antigen of the adult worm of Sj (Sj MAg) in ELISA, furthermore, the characteristics of the scFvs were determined. The soluble scFvs selected were used in assaying sera of acute and chronic patients with Schistosomiasis. Results From the phage displayed antibody library with the content about 1.07×106cfu,two specific positive clones with good reactivity were obtained. Specific soluble anti Sj MAg scFvs were produced from these two clones. The results of Western blot showed that the scFvs of both clones could bind to Sj MAg specifically with a molecular weight about 31kDa. Testing the acute and chronic patients with one of scFvs or two scFvs mixed, the sensitivity is 60%, 36.7%and 75%,56.7%respectively and the overall specificity is 85%～90%. Conclusions The results indicated that a phage displayed library against Sj were constructed successfully and some potential scFvs to detect Sj patients had been isolated.