人源中和性抗汉滩病毒单克隆抗体Fab段基因的获得和表达

运用噬菌体表面表达技术，获得人源和中性抗滩滩病毒汉滩型Ｇ１基因工程单克隆抗体Ｆａｂ段基因及其表达，并同时获得抗汉滩病毒核蛋白的Ｆａｂ抗体。从能综合征出血热疫区恢复期病人抗凝血中分离到的外周淋巴细胞中，提取了部细胞ＲＮＡ。通过ＲＴ－ＰＣＲ方法，用一组人ＩｇＧ Ｆａｂ基因特异性引物，从合成了ｃＤＮＡ中经ＰＣＲ扩增了一组轻链和重链Ｆａｂ段基因，将轻链和重链先后插入噬菌体载体ｐＣｏｍｂ３，ｄｎａｌｆ ｖｆ

GENERATION OF HUMAN NEUTRALIZING MONOCLONAL ANTIBODY Fab FRAGMENTS TO HANTAAN VIRUS BY USING PHAGE DISPLAY TECHNOLOGY

human neutralizing monoclonal antibody Fab fragments to the glycoprotein G1 and a human Fab antibody to the nucleocapsid protein (NP) of hantaan virus have been developed by using phage display technique. The human IgG Fab genes of heavy and light chains were amplified from a HFRS patient in convalescent phase. The combinatorial phage antibody library was prepared by inserting both heavy and light chain Fab genes into phagemid vector pComb3 and followed by the help of phage infection. The library was selected by panning phages which expressed the specific human antibody Fabs on their surface. Total 5 rounds of panning, with alternately using the purified hantaan virus virions and the non-neutralizing Mabs captured glycoproteins, were performed and the specific human Fab fragments to hantaan virus were selected and expressed in bacteria. The protein specificity of the expressed human derived monoclonal antibody Fab fragments were characterized by immune precipitation of radiolabeled hantaan viral proteins, three of the expressed human Fabs showed the G1 specificity and another one was specific to NP of hantaan virus. The specific bindings of the Fab antibodies to hantaan virus were demonstrated by their specific reactions with hantaan virus in ELISA and IFAT. The expressed and purified human Fabs to hantaan virus G1 protein clearly showed the neutralizing activity to HTN virus in plaque reduction neutralization test (PRNT). The results provide poten-tial promise for future use in the prophylaxis or therapy of serious HFRS caused by hantaan virus.