Abstract: Aflatoxin B₁ (AFB₁) has become one of the significant risk factors for human health and food security due to its hypertoxicity and wide distribution in grain. Therefore, it is necessary to detect it. In this study, a novel chromatographic test strip (AuNPs-LFAA) was constructed for the rapid detection of AFB₁ in maize using colloidal gold (AuNPs) and aptamer as the nano-golden probe. Based on the fact that the presence of AFB₁ in the system attenuates the binding of biotin-modified aptamers (Bio-Apt) and thiol-modified complementary chain covalently bound AuNPs (AuNPs-SHcDNA) will be weakened, thusthe color intensity of the test line of the test strip would be reduced.Therefore, how to achieve efficient and sensitive detection of AFB₁ is required. Under the optimal conditions, AuNPs-LFAA detected AFB₁ with a limit of detection (LOD) as low as 0.5 ng/mL, providing a rapid and accurate detection in the range of 0.5-500 ng/mL with high specificity and sensitivity for AFB₁. In summary, AuNPs-LFAA prepared in this study can be an effective method for the detection of AFB₁ in maize and has good application prospects.