[ Objective] To prepare monoclonal antibodies against chicken immunoglobulin G （IgG） and improve the diagnostic level of specific antibodies in chickens. [ Method] Chicken IgG was isolated by saturated ammonium sulf...[ Objective] To prepare monoclonal antibodies against chicken immunoglobulin G （IgG） and improve the diagnostic level of specific antibodies in chickens. [ Method] Chicken IgG was isolated by saturated ammonium sulfate and purified by Sephadex G-200 column chromatography. Then the BALB/c mice were immunized by the chicken IgG, and the spleen cells were fused with mouse myeloma cells SP2/0. Finally, the positive hybridoma cells were screened and detected by indirect enzyme-linked immunosorbent assay （ELISA）. [ Result] Four hybridoma cell strains secre- ting monoclonal antibodies against chicken IgG were obtained and named as C44, C45, C67 and C68, and their ascites titers in indirect ELISA were 1 ： 640 000, 1 ： 320 000, 1 ： 640 000 and 1 ： 80 000, respectively. The monoclonal antibodies secreted by C44 and C45 could recognize light chains of chicken IgG and those secreted by C,67 and C68 could recognize heavy chains of chicken IgG. They all could not recognize IgG from duck, rabbit and swine. Additionally, the Ig type identification results showed that they all belonged to IgGl. [ Conclusion] Four cell strains of obtained hybridoma can stably produce the monoclonal antibodies against chicken IgG.展开更多
[ Objective] To prepare the monoclonal antibody against chicken major histocompatibility complex class II molecules （MHC II）. [ Method ] The prokaryotic expression of the gene fragments of exons 2 -6 encoding alpha ...[ Objective] To prepare the monoclonal antibody against chicken major histocompatibility complex class II molecules （MHC II）. [ Method ] The prokaryotic expression of the gene fragments of exons 2 -6 encoding alpha chain of MHC II and exons 3 -6 encoding beta chain of MHC II were performed based on its protein sequences. After BALB/c mice were immunized with the purified fusion proteins, the mouse spleen cells were fused with mouse myeloma cells SP2/0. Then the positive hybridoma cells were screened and detected by indirect enzyme-linked immunosorbent assay （ELISA）. [ Result] One hybridoma cell strain secreting monoclonal antibody against alpha chain and two strains secreting monoclonal antibody against beta chain were obtained. These three hybridoma cell strains were named as MHC II alpha-4, MHC II betas-2 and MCH II betas-31, respectively. Their titers of ascites in indirect ELISA were 1 ： 256 000, 1 ： 256 000 and 1 ： 1 280 000, respectively. These antibodies could specifically recog- nize MHC II alpha chain or beta chain in western blotting. [ Conclusion] Three obtained hybridoma stains can stably produce the monoclonal antibody against chicken MHC class II molecules.展开更多
利用自动发酵罐培养毕赤酵母,甲醇诱导其分泌表达重组猪IFN-α（r Po IFN-α）;经硫酸铵沉淀、G-25琼脂糖柱脱盐处理及阴离子交换柱和分子筛层析纯化。以纯化r Po IFN-α蛋白作为抗原免疫6周龄BALB/c鼠3次后,取小鼠脾细胞与SP2/0骨髓瘤...利用自动发酵罐培养毕赤酵母,甲醇诱导其分泌表达重组猪IFN-α（r Po IFN-α）;经硫酸铵沉淀、G-25琼脂糖柱脱盐处理及阴离子交换柱和分子筛层析纯化。以纯化r Po IFN-α蛋白作为抗原免疫6周龄BALB/c鼠3次后,取小鼠脾细胞与SP2/0骨髓瘤细胞进行融合,经间接ELISA筛选和3~5次亚克隆,共获得5株均能稳定分泌抗r Po IFN-α单克隆抗体的杂交瘤细胞株。抗体亚类鉴定、特异性和叠加试验结果表明,这5株抗体均属于Ig G1亚类,能与r Po IFN-α产生特异性反应,其抗原结合位点相同或相近。该研究有助于进一步推动r Po IFN-α单克隆抗体在猪免疫学以及猪疫病诊断中的应用。展开更多
基金supported by the National Natural Science Fund (30671537)
文摘[ Objective] To prepare monoclonal antibodies against chicken immunoglobulin G （IgG） and improve the diagnostic level of specific antibodies in chickens. [ Method] Chicken IgG was isolated by saturated ammonium sulfate and purified by Sephadex G-200 column chromatography. Then the BALB/c mice were immunized by the chicken IgG, and the spleen cells were fused with mouse myeloma cells SP2/0. Finally, the positive hybridoma cells were screened and detected by indirect enzyme-linked immunosorbent assay （ELISA）. [ Result] Four hybridoma cell strains secre- ting monoclonal antibodies against chicken IgG were obtained and named as C44, C45, C67 and C68, and their ascites titers in indirect ELISA were 1 ： 640 000, 1 ： 320 000, 1 ： 640 000 and 1 ： 80 000, respectively. The monoclonal antibodies secreted by C44 and C45 could recognize light chains of chicken IgG and those secreted by C,67 and C68 could recognize heavy chains of chicken IgG. They all could not recognize IgG from duck, rabbit and swine. Additionally, the Ig type identification results showed that they all belonged to IgGl. [ Conclusion] Four cell strains of obtained hybridoma can stably produce the monoclonal antibodies against chicken IgG.
基金supported by the National Natural Science Foundation (30671537)
文摘[ Objective] To prepare the monoclonal antibody against chicken major histocompatibility complex class II molecules （MHC II）. [ Method ] The prokaryotic expression of the gene fragments of exons 2 -6 encoding alpha chain of MHC II and exons 3 -6 encoding beta chain of MHC II were performed based on its protein sequences. After BALB/c mice were immunized with the purified fusion proteins, the mouse spleen cells were fused with mouse myeloma cells SP2/0. Then the positive hybridoma cells were screened and detected by indirect enzyme-linked immunosorbent assay （ELISA）. [ Result] One hybridoma cell strain secreting monoclonal antibody against alpha chain and two strains secreting monoclonal antibody against beta chain were obtained. These three hybridoma cell strains were named as MHC II alpha-4, MHC II betas-2 and MCH II betas-31, respectively. Their titers of ascites in indirect ELISA were 1 ： 256 000, 1 ： 256 000 and 1 ： 1 280 000, respectively. These antibodies could specifically recog- nize MHC II alpha chain or beta chain in western blotting. [ Conclusion] Three obtained hybridoma stains can stably produce the monoclonal antibody against chicken MHC class II molecules.
文摘利用自动发酵罐培养毕赤酵母,甲醇诱导其分泌表达重组猪IFN-α（r Po IFN-α）;经硫酸铵沉淀、G-25琼脂糖柱脱盐处理及阴离子交换柱和分子筛层析纯化。以纯化r Po IFN-α蛋白作为抗原免疫6周龄BALB/c鼠3次后,取小鼠脾细胞与SP2/0骨髓瘤细胞进行融合,经间接ELISA筛选和3~5次亚克隆,共获得5株均能稳定分泌抗r Po IFN-α单克隆抗体的杂交瘤细胞株。抗体亚类鉴定、特异性和叠加试验结果表明,这5株抗体均属于Ig G1亚类,能与r Po IFN-α产生特异性反应,其抗原结合位点相同或相近。该研究有助于进一步推动r Po IFN-α单克隆抗体在猪免疫学以及猪疫病诊断中的应用。
