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目的 制备抗溶组织内阿米巴滋养体表面抗原的重组人单克隆抗体Fab段。 方法 提取无症状溶组织内阿米巴包囊携带者外周血淋巴细胞 ,分离总RNA ,行逆转录聚合酶链反应扩增IgG轻链和重链Fd片段 ,并与原核表达质粒连接 ,转化大肠埃希菌构建抗体库。通过抗原夹心膜法、间接荧光抗体试验等方法 ,筛选特异性表达抗溶组织内阿米巴滋养体表面抗原的重组人单克隆抗体Fab段的克隆 ,并纯化其表达产物。 结果 共筛选 3× 10 4个克隆 ,其中一个克隆表达的Fab段识别的抗原表位位于滋养体表面。 结论 运用原核表达系统可制备抗溶组织内阿米巴重组人单克隆抗体Fab段 ,在临床诊断和治疗上均有应用前景
Objective To prepare the Fab fragment of recombinant human monoclonal antibody against Entamoeba histolytica trophozoite surface antigen. Methods Peripheral blood lymphocytes were isolated from asymptomatic Entamoeba histolytica carriers and the total RNA was isolated. The IgG light chain and heavy chain Fd fragments were amplified by reverse transcription polymerase chain reaction (PCR) and ligated with prokaryotic expression plasmids to transform into large intestine Ehrlichia construct antibody library. The recombinant human monoclonal antibody Fab fragment was cloned by antigen sandwich method and indirect fluorescent antibody assay, and the expressed product was purified. Results A total of 3 × 10 4 clones were screened, of which epitopes recognized by one of the cloned Fab fragments were located on the surface of trophozoites. Conclusion The Fab fragment of recombinant human monoclonal antibody against Entamoeba histolytica can be prepared by using prokaryotic expression system and has potential application in clinical diagnosis and treatment