Key Laboratory of Biotechnology Pharmaceutical Engineering of Zhejiang Province, School of Pharmaceutical Science, Wenzhou Medical University, Wenzhou 325000, PR China.
Key Laboratory of Biotechnology Pharmaceutical Engineering of Zhejiang Province, School of Pharmaceutical Science, Wenzhou Medical University, Wenzhou 325000, PR China; Wenzhou Undersun Biotechnology Co., Ltd., Wenzhou 325000, PR China.
J Proteomics. 2017 Aug 8;165:21-25. doi: 10.1016/j.jprot.2017.06.014. Epub 2017 Jun 19.
A highly sensitive method for brief and economical staining of proteins in 1-D and 2-D sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) by 4',5'-Dibromofluorescein (DBF) was developed in this study. Down to 0.025-0.05ng protein could be detected within 10min (only 2 steps) by DBF stain, which is approximately 10-fold more sensitive than those of Eosin Y (EY) and SYPRO Ruby stains, and 20-fold more sensitive than that of imidazole-zinc (IZ) negative stain. In addition, the LC-MS/MS results indicated that the newly developed staining method is compatible with the downstream protein identification. Moreover, the mechanism of DBF stain was investigated by molecular docking.
本研究开发了一种在一维和二维十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）中通过 4',5'-二溴荧光素（DBF）对蛋白质进行短暂、经济染色的高度敏感方法。DBF 染色可在 10min（仅 2 步）内检测到低至 0.025-0.05ng 的蛋白质，其灵敏度大约比曙红 Y（EY）和 SYPRO Ruby 染色高 10 倍，比咪唑-锌（IZ）负染色高 20 倍。此外，LC-MS/MS 结果表明，新开发的染色方法与下游蛋白质鉴定兼容。此外，还通过分子对接研究了 DBF 染色的机制。
