Herein, a ratiometric fluorescence (RF) enzyme-linked immunosorbent assay (RF-ELISA) for sensitive detection of ethyl carbamate (EC) was developed by introducing fluorescent silicon nanoparticles (Si NPs) into the chromogenic substrate system (o-phenylenediamine (OPD)/H₂O₂) of a conventional horseradish peroxidase (HRP)-based ELISA platform to assemble a RF-based signal output system. For this system, the fluorescence of Si NPs at 440 nm (I₄₄₀) was acted as the reference signal which could be efficiently quenched by 2,3-diaminophenazine (DAP), the HRP-catalyzed oxidation product of OPD; Meanwhile, the fluorescence of DAP at 570 nm (I₅₇₀) was served as response signal. Therefore, variation in the amount of HRP labeled secondary antibody bound on the microplate which is associated with antibody-antigen recognition events in conventional HRP-based ELISA could be transferred into a more sensitive RF signal (I₅₇₀/I₄₄₀). On the basis of monoclonal antibody (mAb) which could specifically recognize EC derivative, xanthyl ethyl carbamate (XEC), a Si NPs-based RF-ELISA for EC via a simple pre-analysis derivatization was developed. When detecting the EC content in red wine, this method exhibits a working range from 3.9 to 105.0 μg/L and a limit of detection (LOD) of 2.6 μg/L with excellent specificity, accuracy and reproducibility. The sensitivity is approximately 33-fold higher than that of traditional colorimetric ELISA. The proposed Si NPs-based RF-ELISA is not only highly suitable for screening EC in a large number of samples, but also provides a potential platform for high-throughput and sensitive determination of other analytes for food safety monitoring.

本文中，通过将荧光硅纳米颗粒（Si NPs）引入发色底物系统（o-phenylenediamine（OPD））中，开发了一种用于灵敏检测氨基甲酸乙酯（EC）的比例荧光（RF）酶联免疫吸附测定（RF-ELISA）。 / H ₂ O ₂）的常规辣根过氧化物酶（HRP）-基于ELISA平台，以组装基于RF的信号输出系统。对于该系统，Si NPs在440 nm（I ₄₄₀）处的荧光作为参考信号，可以被HRP催化的OPD氧化产物2,3-二氨基吩嗪（DAP）有效地猝灭。同时，DAP的荧光在570 nm（I ₅₇₀）用作响应信号。因此，与传统的基于HRP的ELISA中的抗体-抗原识别事件相关的微孔板上结合的HRP标记的二抗的数量变化可以转移到更敏感的RF信号中（I ₅₇₀ / I ₄₄₀）。基于可以特异性识别EC衍生物氨基甲酸黄原酸乙酯（XEC）的单克隆抗体（mAb），通过简单的预分析衍生化开发了基于Si NPs的EC的RF-ELISA。当检测红酒中的EC含量时，该方法的工作范围为3.9至105.0μg/ L，检测极限（LOD）为2.6μg/ L，具有出色的特异性，准确性和可重复性。灵敏度比传统比色ELISA高约33倍。所提出的基于Si NPs的RF-ELISA不仅非常适合于筛查大量样品中的EC，而且还为高通量和灵敏地测定其他分析物提供了一个潜在的平台，用于食品安全监控。