Preparative elution of proteins from nitrocellulose membranes after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
Preparative elution of proteins from nitrocellulose membranes after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
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通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分离后，从硝酸纤维素膜上制备性洗脱蛋白质。
DOI：
10.1016/0003-2697(85)90631-1
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发表时间：
1985
影响因子：
2.9
通讯作者：
Montelaro,RC
中科院分区：
文献类型：
--
作者：
Parekh,BS;Mehta,HB;West,MD;Montelaro,RC
对十二烷基硫酸钠（SDS）-聚丙烯酰胺凝胶转移后，从硝酸纤维素膜上制备洗脱蛋白质的各种条件进行了分析和优化。使用吡啶或乙腈洗脱溶剂的洗脱效率最佳，对于含有十二烷基硫酸钠、Triton X-100和脱氧胆酸钠的混合物的缓冲液，为中间体，对于含有任何单一去污剂或离液盐（如尿素或盐酸胍）的缓冲液，洗脱效率可忽略不计。用任何溶剂洗脱的效率也取决于蛋白质的分子量，较小的蛋白质更容易从膜上除去。作为一般程序，蛋白质可通过在5-37 ℃下与40%乙腈或50%吡啶在0.1 m乙酸铵（pH 8.9）中孵育1-3 h从硝酸纤维素膜洗脱。推荐的蛋白质洗脱程序似乎提供了一种快速、简单和有效的方法，可在通过SDS-PAGE分离并转移到硝酸纤维素膜上后从复杂混合物中回收蛋白质。
Various conditions were analyzed and optimized for the preparative elution of proteins from nitrocellulose membranes after transfer from sodium dodecyl sulfate (SDS)-polyacrylamide gels. The efficiency of elution was best using pyridine or acetonitrile elution solvents, intermediate for buffer containing a mixture of sodium dodecyl sulfate, Triton X-100, and sodium deoxycholate, and negligible for buffers containing any single detergent or chaotropic salt, such as urea or guanidine hydrochloride. The efficiency of elution with any solvent also depended on the molecular weight of the protein, smaller proteins being more easily removed from membranes. As a general procedure, proteins may be eluted from nitrocellulose membranes by incubation with either 40% acetonitrile or 50% pyridine in 0.1 m ammonium acetate, pH 8.9, for 1–3 h at 5–37°C. The recommended procedures for protein elution appear to offer a rapid, simple, and efficient means of recovering proteins from complex mixtures after separation by SDS-PAGE and transfer to nitrocellulose membranes.