Aptamer functionalized fluorescent probe for detection of thrombin in human serum via static quenching

Thrombin plays a pivotal role in blood coagulation, wound healing, and tumor metastasis. In this study, we developed a fluorescent probe (Apt15@QDs) by functionalizing quantum dots (QDs) with an aptamer 15 to specifically recognize thrombin. The aptamer Apt15 binds specifically to thrombin by adopting a G-quadruplex conformation, resulting in the formation of a binary complex that quenches the fluorescence of the probe. Analysis using the classical fluorescence quenching equations Stem-Volmer and Lineweaver-Burk revealed that the mechanism of quenching is static in nature. The established method was successfully applied for detecting thrombin in human serum samples. As shown from the results, the established method exhibited high sensitivity, with a limit of detection (LOD) of 8 nM, and possessed a wide linear range within the concentration span of 20 to 200 nM. Additionally, this method circumvented the need for intricate experimental procedures. Furthermore, it offered a short analysis time, obviated the requirement for additional reagents, and proved suitable for high-throughput analysis of biological samples. This study presents a sensitive, straightforward, and rapid approach for detecting thrombin in complex biological matrices.