【摘要】 将合成的非洲猪瘟病毒K205R基因序列插入p ET-28a(+)表达载体,经IPTG诱导表达p ET-28aK205R重组蛋白。用制备的重组抗原免疫BALB/c小鼠,采用化学融合方法融合、筛选,获得了6A4、7F5和7G3三株抗K205R蛋白的杂交瘤细胞株。经鉴定,这3株单克隆抗体(单抗)的亚类均为Ig G1型;单抗的腹水效价分别为1∶51 200、1∶409 600和1∶102 400。将这3株杂交瘤细胞连续培养20代,在第20代仍有很高的分泌抗体的能力,表明其稳定性较好。Western-blot检测结果表明,这3株单抗与K205R重组蛋白有良好的反应性和特异性。上述结果表明,本研究制备的这3株杂交瘤细胞株能够稳定分泌抗体,分泌的抗体特异性高,为非洲猪瘟诊断技术的研究奠定了基础。更多还原

【Abstract】 The synthesized K205 R gene of African swine fever virus(ASFV) was cloned into the p ET-28 a(+)expression vector,and then expressed with IPTG induction.The recombinant K205 R protein was used to immunize BALB/c mice for monoclonal antibody(Mc Ab) preparation,and three hybridoma cell lines(coded6 A4,7 F5 and 7 G3) against the K205 R protein were harvested after chemical fusion and screening.The Ig G1 subtype of the three monoclonal antibodies were identified,and the ascites titers of the McAbs were1∶51 200,1∶409 600 and 1∶102 400,respectively.After continuously cultivation of the three hybridoma cell lines for 20 generations,they were still able to secrete antibodies at a high level,indicating the stability of the cell lines.The three Mc Abs showed good reactivity and specificity with the recombinant K205 R protein in Western-blot.The above-mentioned results showed that the three hybridoma cell lines against recombinant K205 R protein were successfully established for producing Mc Abs,and the Mc Abs had high specificity to K205 R protein of ASFV,which could be useful tool for the development of serological diagnostic tests for African swine fever.更多还原