抗牛碱性成纤维细胞生长因子单克隆抗体的制备及初步鉴定

Preparation and preliminary identification of the monoclonal antibody to bovine bFGF

目的 :制备抗牛碱性成纤维细胞生长因子 (bFGF)单克隆抗体(mAb) ,并鉴定其亚类 ,建立ELISA检测bFGF含量的方法。方法 :应用基因重组的牛bFGF免疫BALB/c小鼠 ,通过细胞融合 ,建立分泌抗bFGFmAb的杂交瘤细胞株。应用免疫沉淀技术鉴定抗bFGFmAb的亚类 ;应用基因重组的牛bFGF免疫青紫蓝兔 ,制备抗bFGF的多抗血清 ;将抗bFGFmAb及兔抗血清用ProteinA亲和层析纯化后 ,建立检测bFGF含量的ELISA方法。结果 :共获得 3株稳定分泌抗bFGFmAb的杂交瘤细胞株 ;它们所分泌的mAb均为IgG1;采用夹心ELISA法检测bFGF的敏感性达ng水平。 结论 :抗bFGFmAb(IgG1)

AIM: To prepare mAb against bovine bFGF and identify their Ig subgroups so as to establish an ELISA for detection of bFGF level. METHODS: BALB/c mice were immunized by recombinant bovine bFGF. Hybridoma cell lines which could stably secret the monoclonal antibodies to bFGF were eytablished by cell fusion technique, and their related characteristics were identified. In addition, polyclonal antibodies to bFGF were prepared by immunization of rabbits with bovine bFGF. The mAb and polyclonal antibodies purified through protein A affinity chromatography were used to develop a sandwich ELISA for detection of bFGF level. RESULTS: Three hybridoma cell lines which could secret the mAbs IgG 1 to bFGF were obtained. The concentration of bFGF could be detected by sandwich ELISA developed with purified mAb and polyclonal antibody at nanogram level. CONCLUSION: mAb and polyclonal antibodies against to bovine bFGF have been prepared successfully, which provide powerful tool for further clinical application and related studies.