Purification and physico-chemical characterization of rabbit tumor necrosis factor.
Purification and physico-chemical characterization of rabbit tumor necrosis factor.
复制标题
兔肿瘤坏死因子的纯化及理化特性。
DOI：
10.4049/jimmunol.125.4.1671
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发表时间：
1980
影响因子：
4.4
通讯作者：
G. Gifford
中科院分区：
文献类型：
--
作者：
M. Ruff;G. Gifford
兔TNF已通过一系列盐沉淀、凝胶过滤、离子交换层析和凝集素亲和层析在SDS-聚丙烯酰胺凝胶电泳（SDS-PAGE）上纯化2000倍至单一种类。TNF活性可以从非变性凝胶系统中恢复，并且已被证明是等电点为5.1的α-球蛋白。M.W.通过SDS-PAGE估计为68，000 d，通过凝胶过滤估计为55，000 d，通过甘油梯度离心估计为52，000 d。TNF活性在6至10的pH范围内是稳定的，并且是相对热稳定的，在70 ℃下1小时不被灭活。TNF活性是链霉蛋白酶敏感的，但相对胰蛋白酶抗性。神经氨酸酶和磷脂酶C治疗没有破坏TNF活性。部分纯化的TNF仍然能够引起易感肿瘤的出血性坏死。粗TNF血清具有3000 U的干扰素滴度，而部分纯化的样品具有<30 U的滴度。
Rabbit TNF has been purified 2000-fold by a series of salt precipitations, gel filtrations, ion exchange chromatography, and lectin affinity chromatography to a single species on SDS-polyacrylamide gel electrophoresis (SDS-PAGE). TNF activity could be recovered from nondenaturing gel systems and has been shown to be an alpha-globulin with an isoelectric point of 5.1. The m.w. was estimated to be 68,000 d by SDS-PAGE, 55,000 by gel filtration, and 52,000 by glycerol gradient centrifugation. TNF activity was stable over the pH range of 6 to 10 and was relatively heat stable, not being inactivated at 70 degrees C for 1 hr. TNF activity was pronase sensitive, but relatively trypsin resistant. Neuraminidase and phospholipase C treatment did not destroy TNF activity. Partially purified TNF was still capable of eliciting hemorrhagic necrosis in susceptible tumors. Crude TNF serum had an interferon titer of 3000 U, whereas the partially purified sample had a titer of <30 U.