禽流感病毒NP蛋白单克隆抗体的研制和鉴定

应用杂交瘤技术,以低致病性禽流感病毒H9N2亚型(Mildly Pathogenic Avian Influenza Virus,MPAIV)F株作为免疫原,制备出3株稳定分泌特异性单克隆抗体的细胞株C2C5、D3B10和C2H8。对单抗的特异性鉴定结果表明:腹水单抗C2C5、D3B10和C2H8的间接ELISA效价分别为5×214、210、5×212,但均无HI效价。3株单抗的免疫球蛋白亚类均为IgG2a。SDS-PAGE和Western-blot分析结果显示:单抗C2H8和D3B10与AIV H9N2和AIV H5N1均有60 kD的蛋白条带反应,而不与其他条带反应,表明这两株单抗特异性地针对AIV H9和AIV H5的共同抗原成分60 kD NP蛋白。而C2C5不与电泳蛋白条带反应,说明该单抗不是针对线性表位的,而可能是针对空间表位的。

Preparation and Identification of Monoclonal Antibodies against NP Protein of Avian Influenza Virus

Taking the F strain of H9N2 subtype of mildly pathogenic avian influenza virus(AIV) as the immunogen,three positive clones(C2C5,D3B10 and C2H8) which could steadily secrete the specific monoclonal antibody were prepared by using hybridoma technique.The results of indirect ELISA and HI tests for the speciality of the above three monoclonal antibodies indicated that the titers of their ascitic fluids were 5×214,210and 5×212 respectively in indirect ELISA test.They were of IgG2a subtype.In SDS-PAGE and Western-blotting analysis,C2H8 and D3B10 could react with NP protein of AIV,while C2C5 could not react with the polypeptide bands of AIV.These results indicated that monoclonal antibodies C2H8 and D3B10 would be useful as a tool for the detection of AIV strains,serological investigation and other epidemiological studies.