Castellanos-Serra Lila, Hardy Eugenio
Division of Physical Chemistry, Center for Genetic Engineering and Biotechnology, Havana, Cuba.
Nat Protoc. 2006;1(3):1544-51. doi: 10.1038/nprot.2006.233.
Here we describe the protocols for negative or reverse detection of proteins, nucleic acids and lipopolysaccharides separated in polyacrylamide electrophoresis gels. These protocols are based on the selective synthesis and precipitation of a white imidazole-zinc complex in the gel, which is absent from those zones where biomolecules are located. These methods are highly sensitive (1-10 ng of biomolecules per band), very cheap as they use inexpensive, common laboratory reagents (imidazole and a Zn II salt), rapid (less than 20 min after gel washing), robust and simple (two steps). Reverse-stained biomolecules are reversibly fixed in the gel. After brief incubation in a zinc chelating agent, biomolecules can be recovered from the gel with the same efficiency as from unstained gels. In consequence, they are procedures of choice for micropreparative applications. References covering typical applications are included.
在此，我们描述了用于聚丙烯酰胺电泳凝胶中蛋白质、核酸和脂多糖阴性或反向检测的方法。这些方法基于在凝胶中选择性合成和沉淀白色咪唑 - 锌络合物，生物分子所在区域不存在该络合物。这些方法高度灵敏（每条带1 - 10 ng生物分子），成本极低，因为使用的是廉价的常见实验室试剂（咪唑和锌盐），速度快（凝胶洗涤后不到20分钟），稳健且操作简单（两步）。反向染色的生物分子在凝胶中可逆固定。在锌螯合剂中短暂孵育后，生物分子可从凝胶中回收，效率与未染色凝胶相同。因此，它们是微制备应用的首选方法。文中包含了典型应用的参考文献。
