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随着标记技术的不断发展,纯化IgG 的需要量不断增加。为了探索一种简单、快速去除绝大多数杂蛋白和白蛋白,而获得免疫球蛋白且不影响其免疫活性的提纯方法,我们对硫酸铵盐析法、DEAE 纤维素离子交换层析法、T 凝胶层析法和批量法等做了比较试验。1 材料和方法1.1 材料(1)免疫腹水:本室用EHFV 的A16株免疫小鼠,以Ehrich 腹水癌诱导产生腹水的方法制备。(2)T 凝胶:丹麦DACO 公司生产,由丹麦哥本哈根大学蛋白质研究室提供。(3)出血热单克隆抗体ELISA 试剂盒:本室生产。1.2 提纯方法(1)硫酸铵盐析法:将腹水2500r/min 离心10min 弃沉淀,用50%和33%饱和硫酸铵沉淀免疫球蛋白,装入透析袋在0.01MpH8.0PB液条件下透析3天。(2)DEAE 纤维素离子交换层析法:文献。(3)T 凝胶层析:将20mlT 凝胶装入16×350mm 柱中,用0.75M 硫酸铵洗柱到最低峰值。等体积的腹水和1.5M 硫酸铵混匀后上样。上样后用0.75M
As labeling technology continues to evolve, the need to purify IgG continues to increase. In order to explore a simple, rapid removal of most of the protein and albumin, and to obtain immunoglobulin and does not affect the immunological activity of the purification method, we ammonium sulfate salting, DEAE cellulose ion exchange chromatography, T Gel chromatography and batch methods such as a comparative test. 1 Materials and methods 1.1 Materials (1) immune ascites: The room with EHFV A16 strains of mice immunized with Ehrich ascites induced ascites produced. (2) T gel: produced by DACO, Denmark, provided by the Protein Laboratory, University of Copenhagen, Denmark. (3) Hemorrhagic fever monoclonal antibody ELISA kit: the production room. 1.2 Purification methods (1) ammonium sulfate salting-out method: the ascites 2500r / min centrifuged 10min discarded precipitate with 50% and 33% saturated ammonium sulfate precipitation immunoglobulin, dialysis bag loaded 0.01MpH8.0PB liquid conditions dialysis 3 day. (2) DEAE cellulose ion exchange chromatography: literature. (3) T-Gel Chromatography: 20 ml of T-gel was loaded into a 16x350 mm column and the column was washed to the lowest peak with 0.75 M ammonium sulfate. Equal volume of ascites and 1.5M ammonium sulfate mixed sample. After loading with 0.75M