Performance Enhancement of Electrochemiluminescence with the Immunosensor Controlled Using Magnetized Masks for the Determination of Epithelial Cancer Biomarker EpCAM

The performance of an electrochemiluminescence (ECL) immunosensor was improved with a particle gradient. SiO₂-coated magnetic beads were adopted as nanocarriers for gradient manipulation and immobilized with the primary antibody. Cadmium telluride quantum dots were coated with a layer of protein G for conjugation and orientation of the secondary antibody as signal labels. ECL immunosensor gradients on the electrode were formed by magnetolithography (ML) with magnetized nickel masks of column and stripe arrays. The immunosensor generally aggregated as an island on the substrate, leading to a decrease of efficiency in the characteristic signals. Stripe arrays of magnetized nickel were designed to generate cylindrical magnetic flux on the substrate to improve the particle manipulation with the gradient. Various gradients of the sandwich-structured immunosensor substantially affected the electrochemical performance. Compared to the gradient-free immunosensor, the gradient of the immunosensor generated by ML using a 3 μm stripe array mask enhanced the ECL intensity ∼2.2 times. The results of quantification of epithelial cell adhesion molecules (EpCAM) with the gradient immunosensor showed a broad linear range (15-420 pg mL^-1), a low limit of detection (5.5 pg mL^-1), and high reliability for EpCAM-spiked serum samples, indicating that the immunosensor gradient substantially enhances the performance of the ECL assay.