Electrochemical Immunosensors for Antibodies to Peanut Allergen Ara h2 Using Gold Nanoparticle-Peptide Films

摘要

Life-threatening allergies to peanuts and tree nuts can be revealed by detecting antibodies (IgEs) to their allergens in patient serum. Herein, we compare several immunosensor-like methodologies for sensitive detection of antibodies to a peptide sequence from the major peanut allergen, Arachis hypogaea 2 (Ara h2). The sensors feature a synthetic peptide layer of the major IgE-binding epitope from Ara h2 attached to a dense gold nanoparticle (AuNP) film on a pyrolytic graphite (PG) electrode. The AuNP-peptide sensor was used to determine model chicken antipeanut antibodies (IgY) in serum. Faradaic and nonfaradaic impedance strategies were compared to amperometric detection. Measurements employed goat antichicken secondary antibodies (Ab_(2)) labeled with horseradish peroxidase (HRP) to bind to IgY on the sensor and provide amplified signals. The best impedimetric sensor configuration featured HPR-catalyzed precipitation of the enzyme product onto the sensor measured by nonfaradaic impedance. This sensor configuration had the best detection limit (DL) of 5 pg mL~(-1) and the best linear range of over 5 orders of magnitude (from 5 pg mL~(-1) to 1 (mu)g mL~(-1)) for IgY antibody in undiluted calf serum. This DL was 100-fold lower than label-free impedimetric immunosensors (0.5 ng mL~(-1)) and 60-fold lower than when using HRP-Ab_(2) in amperometric immunosensors (0.3 ng mL~(-1)).