用单克隆抗体亲和层析法纯化重组人肿瘤坏死因子α

Purification of Recombinant Human Tumor Necrosis Factor Alpha by Affinity Chromatography with Monoclonal Antibodies

用抗rHuTNF-α单抗(G_(11)、D_(12))与溴化氰活化的Sepharose4B偶联制成亲和层析柱,纯化了rHuTNF-α。结果表明,经G_(11)-Sepharose4B柱纯化的TNFα比活性比纯化前提高2.71～4.51倍;经D_(12)-Sepharose4B柱纯化的TNF-α比活性提高2.3～3.4倍。G_(11)-Sepha-rose4B柱的蛋白回收率高于D_(12)-Sepharose4B柱。纯化产物经SDS-PAGE确定达电泳纯,其分子量约为17000道尔顿。

The recombinant human tomur necrosis factor Alpha (rHuTNF-α) was purified with affinity chromatography colunms which were prepared by binding monoclonal antibodies (G_(11), D_(12)) against rHuTNF-α to CNBr-activated Sepharose 4B. It was showed that the biological activitiesof G_(11)-Sepharose 4B-purified rHuTNF-α and D_(12)-Se pharose 4B-purified that increased 2.71~4.51 and 2.3~3.4 fold respectively in comparison with pre-affinity chromatography crude lysates. The rHuTNF-α recovery rate of Gu-Sepharose 4B was higher than that of D_(12)-Sepharose 4B and the purified rHuTNF-α was showed as a single band with a 17KD molecular weight on SDS-PAGE.