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应用SPDP化学偶联法,将天花粉蛋白通过二硫键交联与抗CEA单抗制备成免疫毒素,经Sepbadex G-150凝胶过滤和Blue Sepharose-4B离子梯度交换层析纯化,电泳鉴定显示这一免疫毒素纯度高,偶联蛋白克分子比合理。应用间接免疫荧光染色技术和ELISA法测定结果证明,其与CEA人结肠癌细胞(LoVo)具有特异结合反应,仍保留大部份原抗体活性。具有较好的导向性。体外毒性试验结果表明,免疫毒素对靶细胞(LoVo)有选择性的杀伤作用,50％细胞毒剂量(I.C50)为10~(-9)范围,比单用天花粉蛋白和天花粉蛋白与正常小鼠IgG-偶联物对靶细胞杀伤能力分别增强约150倍和2000倍,而对实验对照细胞(Hela)杀伤无增强作用。
Application of SPDP chemical coupling method, the trichosanthin cross-linked by disulfide and anti-CEA monoclonal antibody prepared immunotoxin, purified by Sepbadex G-150 gel filtration and Blue Sepharose-4B ion exchange chromatography, electrophoresis showed that this A high purity immunotoxin, coupled protein molar ratio is reasonable. The results of indirect immunofluorescence staining and ELISA showed that it has a specific binding reaction with human colon cancer cell line CEA (LoVo), still retaining most of the original antibody activity. Has a good orientation. In vitro toxicity test results showed that immunotoxin selectively killed target cells (LoVo), and the 50% cytotoxic dose (I.C50) was in the range of 10-9, which was higher than that of trichosanthin and trichosanthin The mouse IgG-conjugate enhanced the cytotoxicity of target cells by about 150-fold and 2000-fold, respectively, with no enhancement of killing of experimental control cells (Hela).