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Sensitive, label‐free protein assay using 1‐ethyl‐3‐methylimidazolium tetrafluoroborate‐supported microchip electrophoresis with laser‐induced fluorescence detection
Author(s) -
Xu Yuanhong,
Li Jing,
Wang Erkang
Publication year - 2008
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.200700499
Subject(s) - chemistry , fluorescence , chromatography , dimer , monomer , tetrafluoroborate , detection limit , electrophoresis , trypsin , covalent bond , bovine serum albumin , gel electrophoresis , ionic liquid , biochemistry , organic chemistry , polymer , enzyme , physics , quantum mechanics , catalysis
Based on the dimer–monomer equilibrium movement of the fluorescent dye Pyronin Y (PY), a rapid, simple, highly sensitive, label‐free method for protein detection was developed by microchip electrophoresis with LIF detection. PY formed a nonfluorescent dimer induced by the premicellar aggregation of an anionic surfactant, SDS, however, the fluorescence intensity of the system increased dramatically when proteins such as BSA, bovine hemoglobin, cytochrome c , and trypsin were added to the solution due to the transition of dimer to fluorescent monomer. Furthermore, 1‐ethyl‐3‐methylimidazolium tetrafluoroborate (EMImBF 4 ) instead of PBS was applied as running buffers in microchip electrophoresis. Due to the excellent properties of EMImBF 4 , not only nonspecific protein adsorption was more efficiently suppressed, but also approximately ten‐fold higher fluorescence intensity enhancement was obtained than that using PBS. Under the optimal conditions, detection limits for BSA, bovine hemoglobin, cytochrome c , and trypsin were 1.00×10 −6 , 2×10 −6 , 7×10 −7 , and 5×10 −7 mg/mL, respectively. Thus, without covalent modification of the protein, a protein assay method with high sensitivity was achieved on microchips.