Fluorescence immunoassay based on the enzyme cleaving ss-DNA to regulate the synthesis of histone-ds-poly(AT) templated copper nanoparticles

Abstract

Herein, for the first time we report a novel competitive fluorescence immunoassay for the ultrasensitive detection of aflatoxin B₁ (AFB₁) using histone-ds-poly(AT) templated copper nanoparticles (His-pAT CuNPs) as the fluorescent indicator. In this immunoassay, glucose oxidase (Gox) was used as the carrier of the competing antigen to catalyze the formation of hydrogen peroxide (H₂O₂) from glucose. H₂O₂ was converted to a hydroxyl radical using Fenton's reagent, which further regulated the fluorescence signals of His-pAT CuNPs. Owing to the ultrahigh sensitivity of the ss-DNA to the hydroxyl radical, the proposed fluorescence immunoassay exhibited a favorable dynamic linear detection of AFB₁ ranging from 0.46 pg mL⁻¹ to 400 pg mL⁻¹ with an good half maximal inhibitory concentration and limit of detection of 6.13 and 0.15 pg mL⁻¹, respectively. The intra- and inter-assay showed that the average recoveries for AFB₁ spiked corn samples ranged from 96.87% to 100.73% and 96.67% to 114.92%, respectively. The reliability of this method was further confirmed by adopting ultra-performance liquid chromatography coupled with the fluorescence detector method. In summary, this work offers a novel screening strategy with high sensitivity and robustness for the quantitative detection of mycotoxins or other pollutants for food safety and clinical diagnosis.