重组人粒-巨噬细胞集落刺激因子的分离与纯化

Isolation and Purification of Recombinant Human Granulocyle-macrophage Colony-stimulating Factor

目的：探讨重组人粒－巨噬细胞集落刺激因子的分离，纯化工艺。方法；经破菌溶解包含体，复性等组提纯后，以三步色谱法精制纯化，对终产品进行纯度，生物活性，热原及急性毒性检测。结果；终产品为单一条带，单一峰，生物活性为１．５９×１０＾６－１．８６×１０＾７ＩＵ／ｍｇ热原测定及急性毒性试验符合要求。

Purpose: This study aims at exploring isolation and purification of Recombinant Human Granulocyle- macrophage Colony- stimulating Factor (rhGM-CSF). Methods: After fermentation, centrifuge E.coli cells→ disrupt them by ultrasound→dissolue inclusion bodies→renature→use a three-step chromatography proceduce: hydrophobic chromatography. Ion exchange chromatography and gel filtration→eliminate bacterium. Results: 1. Purity analysis: HPLC method and SDS-PAGE method showed a single peak and a single band corresponding, its purity surpassed 99%; 2. Bioactivity assay: the bioactivity of this product was 1. 59 × 10~7- 1. 86 ×10~7 IU/mg compared with 'Shenbaineng'. 3. Detection of pyrogen and acute toxicity study showed the product was qualified. Conclusion: This method is simple together with good repitition which is suitable for scal-up production.