结核分支杆菌H37Rv株菌体抗原单克隆抗体制备,特性？…

目的 利用单克隆抗体技术分析分支杆菌多肽抗原的共性和个性，建立纯化单克隆抗体的最佳方法。方法 按常规方法制备单克隆抗体。分泌单克隆抗体阳性杂交细胞株的筛选采用ＥＬＩＳＡ法，确认采用免疫印迹法。２４种分支杆菌菌株均生长于改良罗氏培养基上，Ｈ３７Ｒｖ株分别生长于改良罗氏培养基和苏通液体培养基上。单克隆抗体的纯化采用硫酸铵沉淀法、辛酸沉淀法和离子交换法。结果 经筛选获得１４株单克隆抗体杂交细胞瘤。其中，

Production, characterization and purification of monoclonal antibodies against antigens of Mycobacterium tuberculosis H37Rv strain]

OBJECTIVE: Using monoclonal antibody (McAb) technique to analyze polypeptide antigens of Mycobacterium strains and try to find out the best method for purification of McAb. METHODS: McAbs against H37Rv strain were produced by routine procedure. McAb-producing hybridomas were detected by ELISA and immunoblots technique. 24 strains of mycobacterium were grown on Loewenstein-Jensen medium for 1 to 3 weeks at 37 degrees C. H37Rv strain were grown on Loewenstein-Jensen medium or Sauton liquid medium. Ammonium sulfate, caprylic acid precipitation and ion exchange chromatography methods were used to purify McAbs against antigens of H37Rv strain. RESULTS: 14 McAb-produing hybridomas were obtained. C2 McAb reacted with all 24 strains of mycobacterium and 7C12 only reacted with H37Rv, H37Ra, M. bovis, BCG strains, C2 and 7C12 reacted with secreted proteins of H37Rv strain. 40,000/38,000 antigen of H37Rv strain was expressed differently in Loewenstein-Jensen medium and Sauton liquid medium. Relative activity of all McAbs was between 10(-4)-10(-7). The results of purified McAb showed that ammonium sulfate precipitation method could have the highest rate of recovery. Ion exchange chromatography method could have the highest purity. Caprylic acid method can remove albumin effectively and the purification of McAbs in ascific fluid was simple and easy to perform. CONCLUSIONS: Polypeptide antigens of H37Rv against by C2, 7C12 McAb were the secreted proteins. For purification of McAb, the results suggested that ion exchange chromatography method can be applied for analysis and caprylic acid method for a very large volume of ascitic fluid.