Direct Immunoassay for Facile and Sensitive Detection of Small Molecule Aflatoxin B₁ based on Nanobody

Aflatoxin B₁ (AFB₁ ), one of the most toxic mycotoxins, is classified as a group I carcinogen and ubiquitous in various foods and agriproducts. Thus, accurate and sensitive determination of AFB₁ is of great significance to meet the criteria of food safety. Direct detection of AFB₁ is difficult by monoclonal antibody (mAb) with large molecular size (≈150 kD) since the target is too small to produce a detectable signal change. Herein, by combining the electrochemical properties of nanomaterials and the advantages of nanobodies, we developed a direct, highly selective and sensitive electrochemical immunosensor for small molecule detection. The proposed immunosensor had a wide calibration range of 0.01 to 100 ng mL^-1 and a low detection limit of 3.3 pg mL^-1 (S/N=3). Compared with the immunosensor prepared with mAb which was applied in the typical indirect immunoassay, the immunosensor in this work possessed two orders of magnitudes wider linear range and 10-fold more sensitivity. The as-obtained immunosensor was further successfully applied for sensing AFB₁ in real samples. This proposed assay would provide a simple, highly sensitive and selective approach for the direct immunoassay of small molecule AFB₁ , and is extendable to the development of direct immunosensing systems for other small molecules detection by coupling nanocarbon and nanobody.