For real-time and high-sensitivity analysis of low-concentration targets, a sandwich immunoassay using second antibody-second gold nanoparticle (2nd Ab-2nd AuNP) conjugates was combined with fiber-optic localized surface plasmon resonance (FO LSPR). An FO LSPR format was constructed by immobilizing AuNPs on a fiber-optic cross-section for compactness, portability, and ease of handling. In addition, it was combined with a microfluidic system to ensure reproducibility and reliability of measurements. A detection limit of 97.6 fg/mL (148 aM) was obtained for thyroglobulin (Tg) without a sandwich assay. The detection limit was enhanced by approximately 15 times (6.6 fg/mL, 10 aM) when a sandwich strategy was performed with a 2nd Ab-2nd AuNP signal amplifier to further improve the responsivity. Additionally, the good selectivity of the proposed method was confirmed against the unpaired antigen. To evaluate its practical applicability in the field, an FO LSPR biosensor boosted with a sandwich assay using antibody-functionalized AuNPs was applied to detect Tg contained in patient serum, and the results were compared and verified with those of a commercial radioimmunoassay kit. Based on the above results, the signal-enhancing immunoassay with FO LSPR will contribute to the development of optical biosensors for early diagnosis and preventive applications.
Keywords: Antibody-gold nanoparticle conjugate; Biosensor; Localized surface plasmon resonance; Microfluidic chip; Optical fiber; Sandwich immunoassay.
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