论文部分内容阅读
本文构建特异性抗肝癌 (HCC )噬菌体单链抗体 (ScFv )库并观察其靶向性。应用噬菌体表面呈现技术 ,从经HCC细胞免疫的小鼠脾淋巴细胞构建特异性抗HCC噬菌体ScFv库 ,应用HCC细胞和正常肝细胞筛选。制备游离ScFv,以正常肝细胞作为对照 ,应用ELISA、点杂交和免疫组织化学方法以及流式细胞仪 (FCM )检测ScFv与HCC细胞的特异性结合能力。结果显示抗体库库容为 2 14× 10 6 ;核酸序列分析显示ScFv基因结构正确 ;游离ScFv相对分子质量为 3 2 0 0 0 ;ScFv抗HCC效价为 1∶16;点杂交和免疫组织化学技术检测显示HCC细胞被染成棕黄色 ,阴性对照、空白对照未着色 ;FCM检测发现ScFv具有与HCC细胞特异性结合的能力。特异性抗HCC噬菌体ScFv具有与HCC结合的靶向特异性。
A specific anti-hepatoma (HCC) phage single-chain antibody (ScFv) library was constructed and its targeting was observed. Using phage display technology, a specific anti-HCC phage ScFv library was constructed from mouse spleen lymphocytes immunized with HCC cells and screened using HCC cells and normal hepatocytes. Free ScFv was prepared and normal liver cells were used as controls. The specific binding ability of ScFv to HCC cells was detected by ELISA, dot blot and immunohistochemistry, and flow cytometry (FCM). The results showed that the antibody library has a library capacity of 2 14×10 6 ; Nucleic acid sequence analysis showed that the ScFv gene structure is correct; the relative molecular mass of free ScFv is 32000; ScFv anti-HCC titer is 1:16; dot hybridization and immunohistochemistry Detection showed that HCC cells were browned, and negative controls and blank controls were not stained; FCM detection revealed that ScFv had the ability to specifically bind to HCC cells. The specific anti-HCC phage ScFv has a targeting specificity for binding to HCC.