A method for rapid and sensitive negative staining of proteins in SDS‐PAGE using 2′,7′‐dichlorofluorescein |
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| Authors: | Zhongxin Zhu Yewei Huang Jia Sun Zhiwei Chen Litai Jin Yang Wang |
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| Institution: | 1. School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang, P. R. China;2. Wenzhou Undersun Biotechnology Co. Ltd., Wenzhou, Zhejiang, P. R. China;3. Institute of neuroscience, Department of histology and embryology, Wenzhou Medical University, Wenzhou, Zhejiang, P. R. ChinaAdditional corresponding author: Dr. Yang WangE‐mail: |
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| Abstract: | We report here a rapid and sensitive technique for negative visualization of protein in 1D and 2D SDS‐PAGE by using 2′, 7′‐dichlorofluorescein (DCF), which appeared as transparent and colorless bands in an opaque gel matrix background. For DCF stain, down to 0.1–0.2 ng protein could be easily visualized within 7 min by only two steps, and the staining is fourfold more sensitive than that of Eosin Y (EY) negative stain and glutaraldehyde (GA) silver stain, and eightfold more sensitive than that of the commonly used imidazole‐zinc (IZ) negative stain. Furthermore, DCF stain provided good reproducibility, linearity, and MS compatibility compared with those of IZ stain. In addition, the potential staining mechanism was investigated by colorimetric experiment and molecular docking, and the results demonstrated that the interaction between DCF and protein occurs mainly via van der waals force, electrostatic interaction, and hydrogen bonding. |
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| Keywords: | 2′ 7′ ‐Dichlorofluorescein Eosin Y Imidazole‐zinc Negative stain Proteomics |
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