Postelectrophoretic staining of proteins separated by two‐dimensional gel electrophoresis using SYPRO dyes
Postelectrophoretic staining of proteins separated by two‐dimensional gel electrophoresis using SYPRO dyes
复制标题
使用 SYPRO 染料对通过二维凝胶电泳分离的蛋白质进行后电泳染色
作者：
Jun X. Yan;Rachel A. Harry;Carole A. Spibey;M. Dunn
虽然经典银染一直是二维凝胶电泳 (2-D PAGE) 上高灵敏度蛋白质可视化的首选方法，但使用 SYPRO 染料进行电泳后荧光染色的出现对蛋白质组分析中的银染色提出了挑战。后者提供了更高的灵敏度、更高的动态范围并且易于操作。然而，大多数已发表的数据均来自一维凝胶分离分析。在这项工作中，我们重点关注三种市售荧光染料：SYPRO Ruby、SYPRO Orange 和 SYPRO Red（Molecular Probes，尤金，俄勒冈州，美国），并研究了它们在 2D PAGE 上的灵敏度和动态范围。讨论了使用多波长荧光扫描仪对荧光染色可视化的二维蛋白质图谱进行成像，并提供了与银染色分析的详细比较。这些结果证明了使用 SYPRO 染料的优势，与基于一维凝胶电泳的文献一致，并通过二维 PAGE 更真实地了解这些荧光染料的性能。
While the classical silver stain has been the method of choice for high sensitivity protein visualization on two‐dimensional gel electrophoresis (2‐D PAGE), post‐electrophoretic fluorescent staining with the SYPRO group of dyes has emerged to challenge silver staining for proteome analysis. The latter offers improved sensitivity, higher dynamic range and easy handling. However, most of the published data were derived from analysis of 1‐D gel separations. In this work, we have focused on three commercially available fluorescent dyes, SYPRO Ruby, SYPRO Orange and SYPRO Red (Molecular Probes, Eugene, OR, USA) and studied their sensitivity and dynamic range on 2‐D PAGE. The use of a multiwavelength fluorescent scanner to image 2‐D protein profiles visualized with fluorescent staining is discussed, and a detailed comparison with analysis by silver staining is also provided. These results demonstrate the advantages of using SYPRO dyes, which are in agreement with the literature based on 1‐D gel electrophoresis, and give a more realistic understanding of the performance of these fluorescent dyes with 2‐D PAGE.
影响因子：
2.9
作者：
Gatlin, CL;Kleemann, GR;Yates, JR
通讯作者：
Yates, JR