Development of a terminal-fixed aptamer and a label-free colorimetric aptasensor for highly sensitive detection of saxitoxin

Abstract Aptamers show great potentials to be applied for the analysis of small molecules, while the applications of aptamers are now suffering from their weak affinity. Herein, saxitoxin (STX) was chosen as a model target of small molecules. A terminal-fixed anti-STX aptamer (TF-M-30f) was firstly engineered by terminal hybridization. Biolayer interferometry assay showed the terminal-fixed aptamer exhibited a 145-fold higher affinity than the original aptamer. Then, the terminal-fixed aptamer was hybridized with a complementary DNA (cDNA) and fabricated with AuNPs nanozyme to develop a label-free colorimetric aptasensor. In the presence of STX, the aptamer was competitively bound, and the cDNA was released to trigger the enhancement of the peroxidase-like activity of the AuNPs. Computational assays were performed to assist in the rational designs. With the optimal conditions, the developed aptasensor provided a limit of detection as low as 142.3 pM, a linear detection range of 0.1457−37.30 nM, high selectivity, and good recoveries of 98.21–114.1% when analyzing STX in seawater and scallop samples, indicating the high applicability of the engineered aptamer and the developed aptasensor. These innovations can be referred to further develop robust aptamers and aptasensors for small molecules.