亲和层析
色谱法
琼脂糖
化学
大小排阻色谱法
聚丙烯酰胺凝胶电泳
产量(工程)
试剂
生物化学
酶
物理化学
冶金
材料科学
作者
William P. Kolb,Linda M. Kolb,Eckhard R. Podack
出处
期刊:Journal of Immunology
[The American Association of Immunologists]
日期:1979-05-01
卷期号:122 (5): 2103-2111
被引量:89
标识
DOI:10.4049/jimmunol.122.5.2103
摘要
Abstract C1q, a subcomponent of the first component of complement, has been isolated from human serum in fully hemolytically active form by affinity column chromatography and gel filtration with Bio-Gel A-5M. The affinity column was prepared by covalent coupling of purified human IgG to CNBr-activated Sepharose 4B. Final yields of C1q ranged from 25 to 40% with 650- 890-fold purification based on recovery of hemolytic activity. The preparations were free of contaminating serum proteins as judged by SDS-polyacrylamide gel electrophoretic and immunochemical criteria. The final C1q preparations were also devoid of any demonstrable C1q-inhibitor activity. A C1q-depleted reagent (C1qD) was obtained from the nonadsorbed protein containing fractions of the human IgG-Sepharose 4B affinity column and utilized in conjunction with sensitized sheep erythrocytes (EA) for the detection and quantitation of C1q hemolytic activity. Employing optimal quantities of C1qD in the hemolytic assay mixture, the highly purified C1q preparations contained 0.5 to 1 × 1013 effective molecules/mg and 0.5 to 1 × 1012 effective C1q molecules/ml of human serum. This assay would therefore reproducibly detect less than 1 ng of C1q hemolytic activity.
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