Prussian blue-doped CaCO₃ nanoparticle-labeled secondary antibodies for electrochemical immunoassay of interleukin-6 with migraine patients

The sensitive and accurate identification of interleukin-6 (IL-6) in biological fluids is essential for assessing migraine due to its role in different physiological and pathological processes. In this study, we designed a simple and feasible electrochemical immunosensing method for the voltammetric measurement of IL-6. The electrochemical immunosensor was fabricated through covalent conjugation of anti-IL-6 capture antibodies on the glassy carbon electrode with a typical carbodiimide coupling method. Anti-IL-6 secondary antibodies were labeled on the surface of Prussian blue-doped CaCO₃ nanoparticles (PBCaNP) <i>via</i> the epoxy-amino reaction. The assay was carried out with a sandwich-type immunoreaction. In the presence of target IL-6, the analyte was sandwiched between the capture antibody and detection antibody. Thereafter, the carried PBCaNP accompanying the secondary antibody could be detected by using square wave voltammetry (SWV). The voltammetric peak current was dependent on the concentration of target IL-6. Under optimum conditions, the electrochemical immunosensor exhibited good analytical properties, and allowed detection of IL-6 within a wide linear range from 0.1 to 1000 pg mL^-1. The limit of detection was estimated to be 0.078 pg mL^-1 of IL-6 at the 3<i>s</i>_B criterion. An intermediate reproducibility of ≤10.59% was accomplished with batch-to-batch identification, and good anti-interference capacity against other biomolecules was achieved. Importantly, clinical human serum samples obtained from 15 migraine patients were analyzed with the developed electrochemical immunosensors, giving results well-matched with those obtained from the referenced enzyme-linked immunosorbent assay (ELISA) method.