摘要
通过IPTG诱导的大肠埃希菌表达重组牛干扰素-γ融合蛋白(BovIFN-γ),利用GlutathioneSepharose 4 Fast Flow亲和层析柱进行融合蛋白的纯化,纯化后的融合蛋白BovIFN-γ作为抗原免疫Balb/c小鼠,经5次免疫后,取小鼠脾细胞与SP2/0骨髓瘤细胞进行细胞融合,间接ELISA筛选,有限稀释法克隆亚克隆获得分泌BovIFN-γ单克隆抗体的杂交瘤细胞株,并通过ELISA方法对其抗体效价、特异性、亚类及稳定性进行鉴定。获得一株稳定分泌BovIFN-γ单克隆抗体的杂交瘤细胞株A12E9,细胞培养上清和腹水效价可达1∶3 200和1∶160 000。
Recombinant fusion bovine IFN-γ expressed on IPTG-induced E.coli was purified by Glutathione Sepharose 4 Fast Flow affinity column.Balb/c mice were immunized 5 times with purified reccombinant BovIFN-γ protein.Murine myeloma cells were fused with splenocytes of the immunized mice.Positive clones were screened by indirect ELISA and limited dilutions.The main characteristics of screened McAb against BovIFN-γ were further identified by ELISA.Finally,a hybridoma stably secreting specific monoclonal antibody against BovIFN-γ was named A12E9.And the titers of cell culture supernatant and ascitic fluid were 1∶3 200,1∶160 000,respectively.
出处
《动物医学进展》
CSCD
北大核心
2010年第4期53-56,共4页
Progress In Veterinary Medicine
基金
质检公益项目(2007GYJ018)
关键词
牛γ干扰素
单克隆抗体
鉴定
bovine IFN-γ
monoclonal antibody
identification
