Determination of protein tyrosine kinase-7 concentration using electrocatalytic reaction and an aptamer-antibody sandwich assay platform

Abstract

An aptamer-antibody sandwich assay for the voltammetric determination of protein tyrosine kinase-7 (PTK7) concentrations in buffer and human serum solutions was developed. A DNA aptamer specific to PTK7 was covalently immobilized onto gold nanoparticles (AuNPs) modified screen printed carbon electrode (SPCE) surfaces via a self-assembly of mercaptopropionic acid (MPA) followed by a cross linking of the amine modified aptamer. The sequent adsorption of PTK7 and alkaline phosphatase (ALP) conjugated antiPTK7 onto the aptamer surface formed a sandwich complex of PTK7 aptamer/PTK7/antiPTK7-ALP. Following the confirmation of the surface sandwich complex formation with the aptamer, PTK7 and antibody via a series of real-time surface plasmon resonance measurements, the electrocatalytic reaction of surface bound ALP and 4-amino phenyl phosphate (APP) was monitored using cyclic and differential pulse voltammetries. The voltammetric responses linearly increased as a function of the PTK7 concentration ranging from 100 fM to 10 pM with a detectable concentration as low as 100 fM. As a final demonstration, our aptamer-antibody sandwich assay platform was applied to the analysis of PTK7 in biological samples including normal human serum solutions.