交叉免疫电泳法对人抗凝血酶-Ⅲ肝素结合组分的检测  

Crossed immunoelectrophoresis for the heparin-binding fraction detecting of human antithrombin Ⅲ

作　　者：叶生亮[1] 马莉[1] 曹海军[1] 胡吉军 刘凤娟[1] 袁靖 李长清[1] 

机构地区：[1]中国医学科学院北京协和医学院输血研究所,四川成都610052 [2]贵州泰邦生物制品有限公司

出　　处：《中国输血杂志》2015年第1期36-39,共4页Chinese Journal of Blood Transfusion

基　　金：863计划生物和医药技术领域:血浆综合利用及特免球蛋白等新产品开发(2012AA0221903)

摘　　要：目的建立检测人抗凝血酶-Ⅲ(AT-Ⅲ)中肝素结合组分的方法。方法利用火箭电泳法确定AT-Ⅲ抗原抗体比例,以交叉免疫电泳法检测AT-Ⅲ中的肝素结合组分,加热处理方法制备无肝素结合力的AT-Ⅲ样品。结果免疫电泳检测显示沉淀峰清晰,明确分辨出AT-Ⅲ样品中肝素结合组分及非结合组分并计算比例:混合血浆中结合肝素的AT-Ⅲ组分比例约为85.8%,AT-Ⅲ标准品、商业化制品及本实验室自制样品中结合肝素的AT-Ⅲ组分比例均接近100%。AT-Ⅲ样品经热处理(60℃,10h)后,不含枸橼酸钠的样品全部失去肝素结合力,在枸橼酸钠作保护剂的样品中,肝素结合组分占68.5%。结论建立的交叉免疫电泳法能够有效用于人AT-Ⅲ研发、生产时样品中肝素结合组分的检测。Abstract： Objective To establish a method to measure the heparin-binding fraction of human antithrombin Ⅲ （AT- Ⅲ ）. Methods Heparin-hinding fraction is the active native AT-Ⅲ form that has high affinity for heparin. The proportion of heparin-binding fraction must be determined in the quality control of AT-Ⅲ product. Rocket immunoeleetrophoresis （RIE） and crossed immunoelectrophoresis （CIE） were used to measure the heparin-binding fraction of AT-Ⅲ. AT-Ⅲ with low hep-arin binding affinity were prepared using heat treatment. Results Using immunoeleetrophoresis technique, the proportion of the heparin-binding and non-binding fractions of AT-Ⅲ samples could be clearly identified： the fraction of AT-Ⅲ was able to bind to heparin in about 85.8% in pooled plasma. The standard AT-Ⅲ, AT-Ⅲ product and sample from our laboratory were able to approach 100% of binding affinity to heparin. After heat treatment at 60℃ for 10 h, AT-Ⅲ with reduced affinity to heparin was formed. In samples where sodimn citrate were exclu- ded or present, the heparin binding fractions accounted for 0 and 68.5%, respectively. Conclusion The method estab- lished in this study can be used to measure the heparin binding fraction of AT-Ⅲ production effectively.

关 键 词：人抗凝血酶-Ⅲ 肝素结合组分 交叉免疫电泳 火箭电泳 

分 类 号：R392-33[医药卫生—免疫学] R446.1[医药卫生—基础医学]