Thionine-coordinated BCN nanosheets for electrochemical enzyme immunoassay of lipocalin-2 on biofunctionalized carbon-fiber microelectrode

A new electrochemical immune detection protocol on biofunctionalized carbon fiber microelectrode (CFME) was fabricated for ultrasensitive monitoring of lipocalin-2 (LCN2, one typical biomarker of kidney diseases). Two-dimensional boron carbon nitride (BCN) nanosheets with high surface coverage and good conductivity were synthesized and functionalized with the electroactive thionine and peroxidase-conjugated anti-LCN2 detection antibody. The electrochemical immunosensor was constructed through covalent conjugation of anti-LCN2 capture antibody on the CFME. With a sandwich-type immunoreaction, the peroxidase accompanying the immunocomplex exhibited the good electrochemical behaviour with the assistance of redox thionine. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry were utilized for characterization of immunosensor and immunoreaction. Under optimum conditions, BCN-based immunosensing system displayed good electrochemical responses toward target LCN2 within the dynamic range of 0.001-10 ng mL(-1) at a low detection limit of 0.37 pg mL(-1). Meanwhile, the electrochemical immunoassay was also utilized to determine possible cancer biomarkers in human serum with satisfactory specificity. In addition, clinical serum specimens with target LCN2 were investigated on the electrochemical immunosensor using commercial human LCN2 ELISA kit as the reference, and gave well-matched results between two methods.