The authors describe the fabrication of an electrochemical immunosensor for the determination of the activity of protein kinase A (PKA). The method involves (a) electrochemical deposition of gold nanoparticles (AuNPs) on a glassy carbon electrode, (b) PKA-induced catalytic phosphorylation of serine, and (c) the use of phosphoserine antibody and horseradish peroxidase conjugated to IgG on gold nanoparticles (HRP-IgG-AuNPs). The use of AuNPs and HRP-IgG-AuNPs results in large amplification so that the method, at a typical working potential as low as 0.08 V (vs. SCE), has a linear range that extends from 0.1 to 50 activity units per mL, and the detection limit is 0.026 units per mL (at an S/N ratio of 3). The assay is selective (not the least due to a rather low working potential) and well reproducible. It may also be applied to screening for PKA inhibitors and to quantify the PKA activity in human cell lysates.
Microchimica Acta – Springer Journals
Published: Jun 6, 2017
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