Warlow R S, Bernard C C
Psychology Department, La Trobe University, Bundoora, Victoria, Australia.
Electrophoresis. 1990 Jan;11(1):53-60. doi: 10.1002/elps.1150110112.
Optimised silver staining protocols were devised for the detection of membrane proteins in purified form and as a crude mixture. These were adduced in both sodium dodecyl sulphate (SDS) and native polyacrylamide gel electrophoresis and consisted of ethanol-acetic acid-formaldehyde fixation, Coomassie Brilliant Blue prestaining, Rapidfix pretreatment, formaldehyde enhancement and finally ammoniacal silver staining. With these modifications, numerous staining problems of membrane proteins were overcome. These included reduction in background staining, enhanced detection sensitivity in native gels, elimination of negative staining and the avoidance of metallic silver deposition on the gel surface. In overcoming these problems, some factors determining the colour and stainability of membrane proteins in their native state were determined. Both the anionic Coomassie Brilliant Blue dye and SDS detergent improved the sensitivity of silver staining in native gels, and ammoniacal silver was more sensitive than neutral silver, suggesting silver staining to be a charge dependent process.
为检测纯化形式以及粗混合物形式的膜蛋白，设计了优化的银染方案。这些方案应用于十二烷基硫酸钠（SDS）和天然聚丙烯酰胺凝胶电泳中，包括乙醇 - 乙酸 - 甲醛固定、考马斯亮蓝预染、快速固定预处理、甲醛增强，最后进行氨性银染。通过这些改进，克服了膜蛋白染色的诸多问题。这些问题包括背景染色减少、天然凝胶中检测灵敏度提高、阴性染色消除以及避免凝胶表面金属银沉积。在克服这些问题的过程中，确定了一些决定膜蛋白天然状态下颜色和可染性的因素。阴离子考马斯亮蓝染料和SDS去污剂都提高了天然凝胶中银染的灵敏度，并且氨性银比中性银更灵敏，这表明银染是一个依赖电荷的过程。
