Selection of High-Affinity and Selectivity AFB₁ Circular Aptamer for Biosensor Application

Detecting trace amounts of aflatoxin B₁ (AFB₁), one of the most toxic food contaminants, is crucial for efficiently preventing potential health risks. Circular aptamers are promising candidates for bioanalytical applications due to their enhanced biological and structural stability as well as their compatibility with rolling circle amplification (RCA). Herein, we employed a high-efficiency magnetic chain graphene oxide-based SELEX to generate circular aptamers that bind AFB₁ with high affinity and selectivity. Notably, the selected circular aptamer, CAFB₁-A-2, exhibited a significant sequence similarity to the classical AFB₁ linear aptamer but demonstrated a distinct thermodynamic mechanism for binding. The binding of CAFB₁-A-2 was driven by both enthalpy and entropy, whereas the classical linear aptamer exhibited a notable entropy loss that required a greater enthalpy compensation. Utilizing this circular aptamer, we developed a sensitive AFB₁ detection system featuring an RCA-assisted allosteric DNAzyme biosensor with a detection limit (LOD) of 265 pM. Furthermore, the constructed aptasensor successfully detected AFB₁ in spiked rice and corn, achieving LODs of 11 and 9 μg/kg, respectively, within approximately 4 h. This study provides a sensitive and reliable alternative to the development of an aptasensor for AFB₁, holding great potential in the field of food safety detection.