Electrochemical immunosensor for the determination of the cytokine interferon gamma (IFN-γ) in saliva

A simple, fast and sensitive amperometric immunosensing method for the determination of the clinically relevant cytokine interferon gamma (IFN-gamma) in saliva complying the requirements demanded for this kind of sample is reported. The target analyte was sandwiched between a specific capture antibody covalently immobilized on a screen-printed electrode functionalized by the diazonium salt grafting of p-aminobenzoic acid, and a biotinylated detector antibody labeled with a streptavidin-horseradish peroxidase conjugate. The amperometric responses measured at - 0.20 V vs Ag pseudo-reference electrode upon addition of hydrogen peroxide in the presence of hydroquinone as the redox mediator allowed a calibration plot with a linear range between 2.5 and 2000 pg mL(-1) and a low limit of detection (1.6 pg mL(-1)) to be obtained. In addition, a good selectivity against other non-target proteins was achieved. The developed method was validated by analyzing a WHO 1st International Standard for IFN-gamma. In addition, the immunosensor was used for the determination of the endogenous IFN-gamma in saliva with results in excellent agreement with those obtained by a commercial ELISA kit.