用KTA explorer 10蛋白纯化系统分离纯化家蚕类免疫球蛋白

Separation and Purification of Bombyx mori Hemolin with KTA explorer 10 Protein Purification System

类免疫球蛋白(Hemolin)是昆虫的先天性免疫蛋白。为了获取纯度高、具有天然活性的家蚕Hemolin,将被家蚕核型多角体病毒(BmNPV)感染的家蚕蛹进行匀浆处理,离心后对上清液进行DEAE弱阴离子交换层析,进而利用Superdex 200凝胶过滤层析获得家蚕Hemolin的初级纯化产物,经质谱鉴定验证后再利用Superdex 75凝胶过滤层析对初级产物做进一步分离纯化。获得的纯化产物采用SDS-PAGE和Western blot检测到单一的大小约45 kD的蛋白质条带。采用该层析技术与条件能有效制备出纯度较高的具有天然活性的家蚕Hemolin。

Hemolin is a kind of innate immune globulins which have been found in several insect species.To obtain Hemolin with high purity and natural activity from silkworm（Bombyx mori） pupae,Bombyx mori nucleopolyhedrovirus（BmNPV）-infected silkworm pupae were homogenated.After centrifugation,the supernate was purified with DEAE weak anion exchange chromatography and Superdex 200 gel filtration chromatography to yield primary product.After the primary product was verified by mass spectrometry,it was further purified with Superdex 75.A single protein band of about 45 kD was detected from the purified product by SDS-PAGE and Western blot,indicating that it was silkworm Hemolin with high purity.The above results indicate that using these chromatography techniques and conditions can effectively seperate silkworm Hemolin with high purity and natural activity.