Sulfaquinoxaline (SQX) is used extensively in aquaculture and animal husbandry due to its broad antimicrobial spectrum and low cost. However, it is difficult to degrade and there are relevant residues in the aquatic environment, which could be harmful to both the ecological environment and human health. As a new recognition molecule, the aptamer can be recognized with SQX with high affinity and specificity, and the aptamer is no longer adsorbed to AuNPs after binding to SQX, which makes the catalytic effect of AuNPs weakened, an aptasensor for the detection of SQX was successfully constructed. With a linear range of 40-640 ng/mL and a detection limit of 36.95 ng/mL, the aptasensor demonstrated good sensitivity and selectivity. The recoveries of this aptasensor in water samples ranged from 90 to 109.9%, which was quite in line with the high performance liquid chromatography. This indicates that the aptasensor is a useful instrument for detecting SQX in aqueous environmental samples.