A Label-Free Graphene Quantum Dot-Based Fluorescent Aptasensor for Rapid and Sensitive Detection of Leptospira...

Leptospirosis is a neglected tropical disease that is widespread in tropical and subtropical regions. The lack of rapid and sensitive diagnostic tools continues to limit the early identification of the disease. In this study, a graphene quantum dots-based aptasensor was developed for the detection of Leptospira interrogans. Graphene quantum dots were synthesized and used as fluorescence transducers, which are covalently conjugated with cell surface-specific DNA aptamer targeting the pathogen. The synthesized quantum dots exhibited quasi-spherical morphology with particle size ranging from ~ 2.5 nm. Upon target recognition, the GQD-aptamer conjugate exhibited concentrationdependent fluorescence quenching, showing a linear response over the range of 10³-10⁸ CFU/mL (R² = 0.958) and a limit of detection of 10³ CFU mL⁻¹. The assay was completed within 15 min. The sensing platform was further evaluated under biologically relevant conditions, including protein-containing buffer, heat-treated cells, diluted serum, and artificial urine samples. In addition to spectroscopic analysis, dark-field microscopy revealed pronounced aggregation and clustering of Leptospira cells following treatment with the GQD-aptamer conjugate. The combined spectroscopic and microscopic findings support the proposed sensing mechanism and highlight the potential of the developed GQD-aptamer platform as a rapid fluorescence-based aptasensor for Leptospira, with future applicability towards point-of-care diagnostic development.