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来自酸-尿素-曲拉通和十二烷基硫酸钠-聚丙烯酰胺凝胶的组蛋白的蛋白质免疫印迹法。

Western blotting of histones from acid-urea-Triton- and sodium dodecyl sulfate-polyacrylamide gels.

作者信息

Waterborg J H, Harrington R E

出版信息

Anal Biochem. 1987 May 1;162(2):430-4. doi: 10.1016/0003-2697(87)90415-5.

Abstract

We have developed a method for histone transfer from acid-urea-Triton (AUT)-polyacrylamide gels to nitrocellulose filters which prevents the interference of Triton X-100 with the binding of histones to nitrocellulose. Equilibration of AUT gels in 50 mM acetic acid and 0.5% sodium dodecyl sulfate (SDS) allowed displacement of Triton by SDS without loss of band resolution. Electrotransfer of all histone species from treated AUT gels or from equilibrated SDS gels was complete within 1 h in a transfer buffer of Tris-glycine with SDS for increased transfer efficiency and methanol for histone binding. Nitrocellulose with a pore size of 0.2 micron was optimal for histone detection.

摘要

我们开发了一种将组蛋白从酸性尿素-曲拉通(AUT)-聚丙烯酰胺凝胶转移至硝酸纤维素滤膜的方法,该方法可防止曲拉通X-100干扰组蛋白与硝酸纤维素的结合。将AUT凝胶在50 mM乙酸和0.5%十二烷基硫酸钠(SDS)中平衡,可使SDS取代曲拉通,且不损失条带分辨率。在含有SDS以提高转移效率和甲醇以促进组蛋白结合的Tris-甘氨酸转移缓冲液中,所有组蛋白种类从处理过的AUT凝胶或平衡后的SDS凝胶上进行电转移,1小时内即可完成。孔径为0.2微米的硝酸纤维素对组蛋白检测最为理想。

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