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Sequential double fluorescent detections of total proteins and phosphoproteins in SDS ‐ PAGE
Author(s) -
Hwang SunYoung,
Wang Xu,
Cong WeiTao,
Jin LiTai,
Choi JungKap
Publication year - 2014
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201300538
Subject(s) - staining , phosphoprotein , fluorescence , fluorophore , stain , phosvitin , chemistry , chromatography , phosphoproteomics , casein , polyacrylamide gel electrophoresis , microbiology and biotechnology , biochemistry , biology , phosphorylation , protein phosphorylation , protein kinase a , genetics , physics , quantum mechanics , enzyme
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.