Highly sensitive lateral flow immunoassay for aflatoxin B1 detection based on a novel homogeneous enrichment strategy...

Aflatoxin B₁ (AFB₁) is a highly toxic mycotoxin, harms global agriculture and human health. At present, due to the antibody inactivation caused by direct coupling and weak signal output, it is difficult for the traditional lateral flow immunoassays (LFIA) to quickly and accurately detect AFB₁ on the spot. To overcome these limitations, a novel competitive LFIA biosensor was developed utilizing CuS-Au nanocomposites as colorimetric tags and a free nanobody (G8-DIG) as the recognition element. By maintaining the nanobody in a free state rather than immobilizing it, the assay effectively bypasses activity loss while simultaneously amplifying signal output. Under optimal parameters, the proposed sensor quantitatively detects AFB₁ within 8 min, displaying a wide linear range of 0.51-63.3 ng/mL and a low limit of detection (LOD) of 0.26 ng/mL. Furthermore, the method showed excellent selectivity for coexisting mycotoxins and yielded satisfactory recoveries (90.0%-99.6%, CV < 7.4%) in spiked corn samples, which aligned closely with HPLC analysis (95.5%). Overall, the developed immunosensor provides a robust, sensitive, and reliable platform for the rapid point-of-need monitoring of AFB₁ in food.