S/D处理纤维蛋白原制品灭活病毒后去除S/D的方法研究

Method of Removal of Solvent/Detergent Residual in Fibrinogen Product Following Virus Inactivation Treatment by Solvent/Detergent

以低温乙醇法从人血浆中分离的组分Ｉ（Ｃｏｈｎ’ｓ组分Ｉ）为原料，采用有机溶剂／表面活性剂（Ｓ／Ｄ）法进行病毒灭活处理，并用乙醇沉淀法去除Ｓ／Ｄ。实验结果证明：用Ｓ／Ｄ处理的Ｃｏｈｎ’ｓ组分Ｉ经８％的乙醇２次沉淀后，其Ｔｗｅｅｎ８０残留量＜２０μｇ／ｍｌ，ＴＮＢＰ残留量＜１０μｇ／ｍｌ。用醋酸纤维薄膜电泳法测定，纤维蛋白原纯度为９２．７５％，可凝固蛋白含量为９５．５２％，相对回收率为８７．４６％（ｎ＝３）；Ｓ／Ｄ处理前后纤维蛋白原的凝固活力分别为８．０６±０．０８和８．９４±０．７０。ＳＤＳ－ＰＡＧＥ、ＩＥ电泳和分子筛凝胶层析（ＦＰＬＣ）显示无变性的纤维蛋白原存在，提示：乙醇低温条件下沉淀去除Ｓ／Ｄ，不仅能保持纤维蛋白原的生物活性和天然结构基本不变，且可使制品进一步纯化。

The inactivation of virus in Cohns fraction Ⅰ separated from human plasma using cold ethanol method was performed by solvent/detergent(S/D) treatment,and the S/D residual in that fraction was removed by ethanol precipitation.The experimental result showed that Tween 80 residual and TNBP residual in Cohns fraction Ⅰ treated by S/D were <20μg/ml and <10μg/ml,respectively after 8% ethanol precipitation in twice.It was confirmed by cellulose acetate electrophoresis that the purity of fibrinogen was 92.75%,the clottable protein concentration was 95.52% and the relative recovery rate was 87.46%( n =3) and that the clottability of fibrinogen was 8.06±0.08 and 8.94±0.70,respectively before and after S/D treatment.The results of SDS PAGE,immunoelectrophoresis and FPLC demonstrated that there was no denatured fibrinogen,indicating that ethanol precipitation at low temperature for removal of S/D residual in Cohns fraction Ⅰ may not only keep the biological activity and structure of fibrinogen unchanged basically,but also make the product further purified