A lateral flow immunochromatographic assay based on nanobody-oriented coupling strategy for aflatoxin B1 detection

Nonoriented coupling of nanobodies (Nb) to nanomaterials can considerably reduce epitope recognition or induce other dysfunctions, thereby restricting the application of Nb in field detection. Herein, using Avi-tag/streptavidin (Avi/SA)-oriented coupling strategy, we developed a lateral-flow immunochromatographic assay (LFIA) based on Nb-Avi/SA@quantum dots (QDs) probes (Nb-LFIA) for Aflatoxin B1 (AFB1) detection. Compared with nonoriented coupled Nb@QD probes, the detection sensitivity and stability of Nb-Avi/SA@QD probes were greatly retained. Furthermore, Nb showed higher tolerance to the extreme detection environments compared with monoclonal antibodies in traditional enzyme-linked immunosorbent assay and LFIA. After optimization, the limit of detection of Nb-LFIA was 0.095 ng mL−1, half-maximal inhibitory concentration was 0.85 ng mL−1, and the visual cut-off level was 1.25 ng mL−1. Nb-Avi/SA@gold nanoparticle probes were assessed for the wide applicability of the Avi/SA strategy, and they showed excellent performance (coefficient of determination = 0.999). Finally, the analysis performance of probes was assessed in oat samples, revealing a recovery rate of 88.8–116.7%, which was consistent with the results of high-pressure liquid chromatography coupled with tandem mass spectrometry. Altogether, our study provides a convenient and sensitive tool for AFB1 detection, and presents a viable path for efficient Nb application.