Graphdiyne oxide as a high-capacity biointerface for multiplexed electrochemical immunosensing of neutrophil biomarkers.

The rapid and reliable quantification of inflammatory biomarkers is highly desirable for clinical diagnostics and point-of-care testing. In this work, we have developed a dual electrochemical immunosensing platform based on graphdiyne oxide (GDYO) for simultaneously determining the neutrophil activation biomarkers CD63 and CD66b. GDYO was integrated onto screen-printed carbon electrodes to create a high-capacity biointerface with an abundance of reactive sites for antibody immobilization and efficient signal transduction. The analytical approach relied on a sandwich immunoassay configuration that employed horseradish peroxidase (HRP)-labeled secondary antibodies. Amperometric detection was performed using the hydroquinone/hydrogen peroxide redox system. Under optimized conditions, the platform enabled the simultaneous quantification of both analytes within a concentration range of 0.005-100 ng·mL-1, achieving limits of detection (LOD) of 0.1 and 0.6 pg·mL-1 for CD63 and CD66b, respectively. The immunosensor exhibited satisfactory reproducibility, storage stability, and consistent analytical performance. Its practical applicability was demonstrated by analyzing serum samples from healthy individuals and patients diagnosed with chronic obstructive pulmonary disease (COPD). The proposed GDYO-based electrochemical platform is a promising strategy for the sensitive determination of neutrophil-associated biomarkers in decentralized clinical testing.