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在未密封的聚乙烯袋中长期无缓冲湿凝胶储存。

Long-term, buffer-less, wet gel storage in non-sealed polyethylene bags.

作者信息

Kurien Biji T, Scofield R Hal

机构信息

Department of Veterans Affairs, University of Oklahoma Health Sciences Center, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA.

出版信息

Methods Mol Biol. 2012;869:437-43. doi: 10.1007/978-1-61779-821-4_37.

Abstract

Electrophoresed gels are normally fixed, stained, destained, and dried. Drying is normally carried out with commercial gel dryers or by drying between two cellophane sheets held together by two acrylic frames. Here, we report that stained and destained gels (7.5, 10, or 15%, denaturing or native gels; 0.4 or 1.5 mm in thickness) could be stored wet, unsealed, and without any storage buffer for several months at room temperature within flexible polyethylene bags without significant shrinking or protein diffusion. The gel remains hydrated because of the de facto sealing achieved by the polyethylene sheets (PS) adhering airtightly to the gel on either sides. The microsaturated environment generated by the thin film of water molecules trapped between the gel and the PS, along with the nonporous nature of the PS, apparently protects the gel from cracking as well as shrinking significantly. The intensity of stained proteins increased during storage probably from the slight gel shrinkage observed. Wet gel storage is useful (a) when low abundance protein spots from multiple two-dimensional electrophoresis gels have to be excised for in-gel tryptic digestion or electroelution and (b) for wet gel autoradiography. In addition to avoiding dryer contamination and saving drying time, these bags prevent the moist gel from sticking to X-ray film. Such storage could also prove useful for electrophoretic transfer of fixed and stained gels.

摘要

电泳后的凝胶通常要进行固定、染色、脱色和干燥处理。干燥通常使用商用凝胶干燥器进行,或者通过夹在两个丙烯酸框架之间的两张玻璃纸之间晾干。在此,我们报告,染色和脱色后的凝胶(7.5%、10%或15%,变性或非变性凝胶;厚度为0.4或1.5毫米)可以在室温下,在柔性聚乙烯袋中无密封、不添加任何储存缓冲液的情况下湿存数月,且不会显著收缩或出现蛋白质扩散现象。由于聚乙烯片材(PS)紧密贴附在凝胶两侧实现了事实上的密封,凝胶得以保持水合状态。凝胶与PS之间截留的水分子薄膜形成的微饱和环境,以及PS的无孔性质,显然保护了凝胶不发生开裂以及显著收缩。储存期间染色蛋白质的强度增加,可能是由于观察到凝胶略有收缩。湿凝胶储存很有用:(a)当必须从多个二维电泳凝胶中切下低丰度蛋白质斑点用于胶内胰蛋白酶消化或电洗脱时;(b)用于湿凝胶放射自显影。除了避免干燥器污染和节省干燥时间外,这些袋子还能防止湿凝胶粘在X射线胶片上。这种储存对于固定和染色凝胶的电泳转移也可能有用。

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