Preparation of monoclonal antibody against pA104R protein of African swine fever virus by semi-solid culture method
In order to prepare the monoclonal antibodies against African swine fever virus(ASFV)rapidly and effi-ciently,the recombinant pA104R protein of ASFV was systematically expressed and purified by Escherichia coli in this stud-y.The recombinant pA104R protein of ASFV was used as antigen to compare the two immune strategies of CpG ODN combined with aluminum hydroxide adjuvant and conven-tional Freund's adjuvant,respectively,and the efficiency of semi-solid culture method and conventional limited dilu-tion method in the preparation of monoclonal antibody a-gainst ASFV pA104R was compared.The results showed that the recombinant pA104R soluble protein of ASFV with a relative molecular weight of 3.5×104 was obtained,and the fu-sion requirement was achieved on the 21st day after the mice were immunized with CpG ODN and aluminum hydroxide adju-vant.Compared with routine Freund's adjuvant immunization and recombinant pA104R protein,CpG ODN combined with aluminum hydroxide adjuvant and semi-solid culture method in this experiment saved 14 days.The semi-solid culture meth-od shortened the test period of monoclonal screening by 28 days compared with the limited dilution method,and saved a lot of subclonal workload.Five positive hybridoma cell lines were obtained by semi-solid culture method,and three high titer hybridoma cell lines(9A4,9H6,11F5)were selected for identification.The heavy chain was IgG,and the light chain was KAPPA.The titers of three purified monoclonal antibodies against pA104 protein and whole virus protein were 1∶160 000-1∶320 000 and 1∶200-1∶400,respectively.In this study,monoclonal antibodies were screened by CpG combined with aluminum hydroxide adjuvant and semi-solid culture method,providing a new rapid and efficient strategy for monoclonal an-tibody preparation.
African swine fever viruspA104 proteinmonoclonal antibodysemi-solid medium method
万方数据
维普
