辛酸沉淀结合阴离子交换层析纯化马IgG的研究  

Fractionation of equine IgG using caprylic acid precipitation in combination with anion exchange chromatography

作　　者：杨俊杰[1] 段丽娟[1] 金学敏[1] 孙烈英[1] 任正祥 孟祥玉[1] 张金[1] 高建军[1] 

机构地区：[1]兰州生物制品研究所有限责任公司 甘肃省疫苗工程技术研究中心,甘肃兰州730046

出　　处：《微生物学免疫学进展》2015年第1期17-20,共4页Progress In Microbiology and Immunology

摘　　要：目的以辛酸沉淀结合离子交换层析纯化破伤风抗毒素马免疫血浆,获得高质量的马IgG,为抗毒素F(ab')2的制备奠定基础。方法通过对辛酸沉淀马血浆过程中的pH、辛酸浓度以及血浆稀释倍数的实验设计(DoE),研究不同条件对IgG纯度、效价、比活性、浊度以及过滤速度的影响,确定各工艺参数的可操作区间,并结合Capto DEAE阴离子交换层析以流穿模式进一步纯化IgG。结果马血浆经一步辛酸沉淀获得的IgG纯度(SDSPAGE)大于92%、分子排阻色谱(SEC)纯度大于95%,比活性较血浆提高2.14±0.29倍;辛酸沉淀后的IgG样品经阴离子交换层析,可有效去除聚合体以及小分子杂质,将纯度提高至95%(SDS-PAGE)和98%(SEC)。结论马血浆经辛酸沉淀和阴离子交换层析可获得高纯度、高比活的IgG。Objective Fractionation of high quality IgG from equine plasma by using caprylic acid （CA） precipitation in combination with anion exchange chromatography. To provide a basis for the preparation of antitoxin F （ ab＇ ） 2. Methods Optimization of CA precipitation of equine plasma for IgG preparation was achieved by regression analysis of the responses of three highly significant factors assayed by design of experiment （ DoE ）, the factors studied were CA concentration, pH and plasma dilution, and their response was assessed in terms of purity, potency, activity, turbidity and filtration speed. The results evidenced the operational range of three variables in the process. Then, anion exchange chromatography was adopted for further purification by a flow through mode. Results The preparation obtained by one step CA precipitation contains more than 92% of equine IgG（SDS-PAGE）, and SEC analysis showed the purity was more than 95%. The activity of purified IgG was 2.14±0.29-fold higher than that of equine plasma. The supernatant from CA precipitation was then subjected to a Capto DEAE column chromatography for the elimination of aggregates and some low molecular components, and this lead to the purity further reached to 95% （ SDS-PAGE ） and 98.79% （ SEC ）, respectively. Conclusion Equine IgG obtained by CA precipitation in combination with anion exchange chromatography presented high purity and activity.

关 键 词：破伤风抗毒素 辛酸 离子交换层析 实验设计(DoE) 

分 类 号：S852.4[农业科学—基础兽医学]