Electrochemical detection of cardiac biomarker myoglobin using polyphenol as imprinted polymer receptor.

ABSTRACT

An electrochemical biosensor was developed by merging the features of Molecular Imprinting technique and Screen-Printed Electrode (SPE) for the simple and fast screening of cardiac biomarker myoglobin (Myo) in point-of-care (POC). The MIP artificial receptor for Myo was prepared by electrooxidative polymerization of phenol (Ph) on a AuSPE in the presence of Myo as template molecule. The choice of the most effective protein extraction procedure from the various extraction methods tested (mildly acidic/basic solutions, pure/mixed organic solvents, solutions containing surfactants and enzymatic digestion methods), and the optimization of the thickness of the polymer film was carefully undertaken in order to improve binding characteristics of Myo to the imprinted polymer receptor and increase the sensitivity of the MIP biosensor. The film thickness was optimized by adjusting scan rate and the number of cycles during cyclic voltammetric electropolymerization of Ph. The thickness of the polyphenol nanocoating of only few nanometres (∼4.4 nm), and similar to the protein diameter, brought in significant improvements in terms of sensor sensitivity. The binding affinity of MIP receptor film was estimated by fitting the experimental data to Freundlich isotherm and a ∼8 fold increase in the binding affinity of Myo to the imprinted polymer (KF = 0.119 ± 0.002 ng-1 mL) when compared to the non-imprinted polymer (KF = 0.015 ± 0.002 ng-1 mL) which demonstrated excellent (re)binding affinity for the imprinted protein. The incubation of the Myo MIP receptor modified electrode with increasing concentration of protein (from 0.001 ng mL-1 to 100 µg mL-1) resulted in a decrease of the ferro/ferricyanide redox current. LODs of 2.1 and 14 pg mL-1 were obtained from calibration curves built in neutral buffer and diluted artificial serum, respectively, using SWV technique, enabling the detection of the protein biomarker at clinically relevant levels. The prepared MIP biosensor was applied to the determination of Myo spiked serum samples with satisfactory results.