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lzangelfish

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[交流] [交流求助]酶的纯化

各位虫子好,我做的是大豆脂肪氧和酶LOX1的分离纯化
此酶的分子量大约是102KDa,等电点约在5.70
按文献来说应采用DEAE Sephadex A 50 在0.02M,pH6.8的PBS下进进行分离纯化。
而后用0.02-0.17 0.17-0.23的PBS分部梯度洗脱
我想问一下各位虫子DEAE Sephadex A 50 的上样量一般柱体积的多少?我柱体积约90ml,应上多少mg蛋白?什么浓度?
另外在0.02-0.17时流速为30ml/H.酶应在0.2M左右出峰,我是否应在第二部梯度洗脱时放慢流速?
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