A split aptamer-based biosensor for visual detection of leuco-malachite green

Leuco-malachite green (LMG), a metabolite of malachite green (MG), exhibits stronger bioaccumulation and higher toxicity to humans. Therefore, development of a rapid and sensitive detection method for LMG is of great significance. In this study, LMG-specific split aptamers (SPALMG) with a dissociation constant (Kd) of 1.538 ± 0.1261 μM and enhanced affinity compared to the parent aptamer are generated using A5b-12C, a high-affinity variant, as the parent aptamer for LMG detection. Subsequently, a colorimetric aptasensor is developed, employing thiol-modified SPAlmg as probes and gold nanoparticles (AuNPs) as signal indicators. In the presence of LMG, the SPAlmg preferentially bind to LMG to form a ternary complex, causing the AuNPs to aggregate and induce a color change from red to blue. The sensor exhibits high sensitivity and specificity, with a limit of detection (LOD) of 46 nM, a linear range of 1–7 μM, and good recoveries of 96–113%. Compared with other studies, a more excellent LOD is achieved. This work provides a novel affinity probe that holds great promise for the development of sandwich-type sensors for LMG detection.