Hwang Sun-Young, Wang Xu, Cong Wei-Tao, Jin Li-Tai, Choi Jung-Kap
College of Pharmacy & Research Institute of Drug Development, Chonnam National University, Gwangju, South Korea.
Electrophoresis. 2014 Apr;35(8):1089-98. doi: 10.1002/elps.201300538. Epub 2014 Mar 20.
A fluorescent staining technique, using selective chelation with fluorophore and metal ion to the phosphate groups of phosphoproteins in SDS-PAGE is described. As a fluorescent dye and a metal ion, Fura 2 pentapotassium salt and Al(3+) were employed, respectively. The staining method, Fura 2 stain, has sensitivities of 16-32 ng of α-casein and β-casein, 62 ng of ovalbumin, phosvitin, and κ-casein using an ultraviolet transilluminator. Furthermore, Fura 2 stain is able to carry out continuative double detection of total proteins and phosphoproteins on the same gel within 3.5 h. Consequently, selective phosphoprotein and total protein detections could be obtained without other poststaining. Considering the low cost, simplicity, and speed, Fura 2 staining may provide great practicalities in routine phosphoproteomics research.
本文描述了一种荧光染色技术，该技术利用荧光团与金属离子对SDS-PAGE中磷蛋白磷酸基团的选择性螯合作用。分别使用Fura 2五钾盐作为荧光染料和Al(3+)作为金属离子。这种染色方法，即Fura 2染色法，使用紫外透照仪时，对α-酪蛋白和β-酪蛋白的灵敏度为16 - 32 ng，对卵清蛋白、卵黄高磷蛋白和κ-酪蛋白的灵敏度为62 ng。此外，Fura 2染色能够在3.5小时内在同一凝胶上对总蛋白和磷蛋白进行连续双重检测。因此，无需其他染色后处理即可实现磷蛋白和总蛋白的选择性检测。考虑到成本低、操作简单和速度快，Fura 2染色在常规磷酸化蛋白质组学研究中可能具有很大的实用性。
