Farci Domenica, Kirkpatrick Joanna, Piano Dario
Laboratory of Plant Physiology and Photobiology, Department of Life and Environmental Sciences, University of Cagliari, Cagliari, Italy.
Department of Molecular Sensory Systems, Center of Advanced European Studies and Research (Caesar), Bonn, Germany.
Electrophoresis. 2017 Feb;38(3-4):441-446. doi: 10.1002/elps.201600389. Epub 2016 Nov 22.
We report a fast and sensitive procedure for blue native PAGE staining, in which the conventional staining step with CBB is avoided. After running, a short exposure to a mix of polar protic solvents (ethanol and acetic acid) leads to a fast and selective removal of the dye from the migration front and a specific binding to the protein bands, while the rest undergo a selective and complete background removal, leading to an intense contrast. This single-step staining-destaining technique is useful in protein samples that bind colored cofactors such as photosystems, which can be selectively discerned by their characteristic green color. After the staining of such samples, the green color persists, while the other unpigmented protein complexes and the molecular standard remain CBB stained, creating a useful reference system for the assignment of the bands. The advantages and chemical basis of this staining procedure are discussed.
我们报告了一种用于蓝色天然聚丙烯酰胺凝胶电泳染色的快速且灵敏的方法，该方法避免了使用考马斯亮蓝（CBB）的传统染色步骤。电泳结束后，短暂暴露于极性质子溶剂（乙醇和乙酸）的混合液中，可快速且选择性地将染料从迁移前沿去除，并使其特异性结合到蛋白条带上，而其余部分则经历选择性且完全的背景去除，从而产生强烈的对比度。这种单步染色-脱色技术适用于结合有色辅因子的蛋白质样品，如光系统，其可通过特征性的绿色被选择性识别。对此类样品进行染色后，绿色会保留，而其他无色素的蛋白质复合物和分子标准品仍被CBB染色，从而创建了一个用于条带归属的有用参考系统。本文讨论了该染色方法的优点和化学原理。
