Label-free electrochemical impedance detection of kinase and phosphatase activities using carbon nanofiber nanoelectrode arrays

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Show simple item record Li, Yi Fen Syed, Lateef Liu, Jianwei Hua, Duy H. Li, Jun 2012-09-26T14:30:00Z 2012-09-26T14:30:00Z 2012-09-26
dc.description.abstract We demonstrate the feasibility of a label-free electrochemical method to detect the kinetics of phosphorylation and dephosphorylation of surface-attached peptides catalyzed by kinase and phosphatase, respectively. The peptides with a sequence specific to c-Src tyrosine kinase and protein tyrosine phosphatase 1B (PTP1B) were first validated with ELISA-based protein tyrosine kinase assay and then functionalized on vertically aligned carbon nanofiber (VACNF) nanoelectrode arrays (NEAs). Real-time electrochemical impedance spectroscopy (REIS) measurements showed reversible impedance changes upon the addition of c-Src kinase and PTP1B phosphatase. Only a small and unreliable impedance variation was observed during the peptide phosphorylation, but a large and fast impedance decrease was observed during the peptide dephosphorylation at different PTP1B concentrations. The REIS data of dephosphorylation displayed a well-defined exponential decay following the Michaelis–Menten heterogeneous enzymatic model with a specific constant, k[subscript]c[subscript]a[subscript]t/K[subscript]m, of (2.1 ± 0.1) × 10[superscript]7 M[superscript]−[superscript]1 s[superscript]−[superscript]1. Consistent values of the specific constant was measured at PTP1B concentration varying from 1.2 to 2.4 nM with the corresponding electrochemical signal decay constant varying from 38.5 to 19.1 s. This electrochemical method can be potentially used as a label-free method for profiling enzyme activities in fast reactions. en_US
dc.relation.uri en_US
dc.subject Real-time electrochemical impedance spectroscopy en_US
dc.subject Phosphorylation en_US
dc.subject Dephosphorylation en_US
dc.subject Heterogeneous enzyme kinetics en_US
dc.subject Vertically aligned carbon nanofibers en_US
dc.subject c-Src tyrosine kinase en_US
dc.subject Protein tyrosine phosphatase 1B en_US
dc.subject Nanoelectrode array en_US
dc.title Label-free electrochemical impedance detection of kinase and phosphatase activities using carbon nanofiber nanoelectrode arrays en_US
dc.type Article (author version) en_US 2012 en_US
dc.citation.doi doi:10.1016/j.aca.2012.07.027 en_US
dc.citation.epage 53 en_US
dc.citation.jtitle Analytica Chimica Acta en_US
dc.citation.spage 45 en_US
dc.citation.volume 744 en_US
dc.contributor.authoreid duy en_US
dc.contributor.authoreid yifenli en_US
dc.contributor.authoreid lateefs en_US
dc.contributor.authoreid junli en_US

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