离子对染色体系细菌蛋白染色效果分析

Application of ion pair staining system in process of bacterial protein staining

目的评价蛋白离子对染色体系（使用2种带反向电荷物质）对细菌蛋白染色效果,并比较其与同类试剂的异同。方法纯化扩增金黄色葡萄球菌、副溶血弧菌等菌株,提取菌体蛋白,定量,4倍梯度稀释,十二烷基磺酸钠（SDS）聚丙烯酰胺凝胶电泳。分别用离子对染色体系与商品化进口试剂盒对凝胶进行染料染色和银染,对比分析染色效果。结果离子对染料最低可使0.16μg细菌蛋白显出条带,与进口染色试剂盒接近,但背景更干净,条带更清晰;离子对银染与国外进口试剂银染比较,操作更加简便,蛋白条带着色迅速而清晰,且可耐受较长时间显色（10~20 m in）;离子对染色体系中染料是银染的敏化剂,染料染色后再行银染与直接银染效果一致,且可省去敏化过程。结论离子对染色体系的染料染色与银染有很好的兼容性,染色结果重复性明显提高,更贴近蛋白分离染色检测的需要。

Objective To evaluate a new protein ion pair staining system（adopting two kinds of material with reverse charge）for bacterial protein staining,and to compare its difference with similar foreign commercial staining kit.Methods Bacterial proteome of strains of Staphylococcus aureus and Vibrio parahaemolyticus were extracted with thermo solution and quantified with the RC DC kit and diluted with 4-folds gradient,and then followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.The gels were treated with dye staining and re-silver staining by contrast ion-pair system and imported commercial staining kits.Results The new type of dyes resulted in a more clear and obvious lane than that of foreign commercial staining kit with a bacterial protein limitation as low as 0.16 μg.Compared to foreign commercial staining kit,the operation for ion pairs silver staining system was more simple and the background was clearer;the display of bands was quick and clear.With ion pairs silver staining,the gels could withstand a longer period staining.In the process of ion pairs staining,the dye is a sensitizing agent for silver staining.The results of re-silver staining after dyeing were similar to the direct silver staining,and the process of sensitizing could be omited.Conclusion The dye used ion pair staining system has good compatibility with silver staining and is more suitable for the requirement of bacterial proteomics.