Gold Nanoparticle/Nanobody Immunochromatographic Strip for Detection of 19-Nortestosterone

19-nortestosterone (19-NT) is recognized as a highly effective anabolic steroid that is banned in animal products because of its negative impact on human health. Despite legal prohibitions, 19-NT is still abused, driven by its commercial benefits. In this study, a gold nanoparticle immunochromatographic assay (GICA) utilizing nanobodies (Nb) has been developed for the efficient, rapid, and sensitive detection of 19-NT. Specifically, anti-19-NT-Nb2F7 coupled gold nanoparticles and optimized a variety of detection conditions for GICA, including pH, concentration of Nb2F7, and amount of probe. Under optimal conditions, the Nb2F7-based GICA could rapidly and sensitively detect 19-NT within 10 min, with a limit of detection (LOD) as low as 0.08 ng/mL. Moreover, this method exhibited good recovery and stability for detecting 19-NT in pig urine and bovine urine, which was correlated well with standard UPLC-MS/MS. Overall, this Nb2F7-based GICA is well-suited for on-site screening of 19-NT in animal urine products, providing a rapid, cost-effective, and user-friendly alternative to traditional assay.