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利用GM1偶联活化的Sephadex G50或扰CT IgG偶联活化的Sepharose 4B,我们已成功地建立了两种亲和层析纯化CT和CT-B的方法,获得了纯化产品。受体亲和柱对CT-B和CT的分离效率分别为71.6％或48.3％,抗体亲和柱分别为50％或51.7％。纯化产物经SDSPAGE、PAGE分析表明均达到亲和层析电泳纯级。GM1-ELISA、CHO细胞测毒加间接免疫荧光检测、家兔肠段结扎试验和家兔免疫保护试验结果提示纯化产物具有良好的生物学活性。这种有效而简单的方法可推广应用于CT或CT相关毒素的纯化。
We have successfully established two methods for purifying CT and CT-B by affinity chromatography using GM1 coupled with activated Sephadex G50 or CT-conjugated IgG-activated Sepharose 4B to obtain the purified product. The separation efficiency of CT-B and CT for receptor affinity column was 71.6% or 48.3%, respectively, and the affinity columns were 50% or 51.7% respectively. The purified product was SDSPAGE, PAGE analysis showed that all reached affinity chromatography electrophoresis grade. GM1-ELISA, CHO cell assay plus indirect immunofluorescence assay, rabbit intestine ligation test and rabbit immunoprotection assay indicated that the purified product had good biological activity. This effective and simple method can be extended to purify CT or CT related toxins.