Advancement of doxorubicin monitoring with a DNA fragmentation strategy for SYBR Green I-based aptamer biosensors

Doxorubicin (DOX), a widely utilized chemotherapeutic antibiotic, increasingly threatens environmental and public health due to its persistence in marine ecosystems, bioaccumulation in organisms, and contamination of agricultural systems. In response to the urgent demand for effective DOX detection, we have developed a label-free fluorescent aptamer biosensor (FAB) that employs SYBR Green I (SG I)-mediated signal amplification. A key innovation in this design is the strategic fragmentation of the 41-mer complementary strand (CP) of the DOX aptamer into three 11-mer fragments. It significantly mitigates nonspecific SG I binding interference caused by the longer ssDNA CP, enhancing the fluorescence signal ratio (F dsDNA /F ssDNA ) from 4.6-fold to 12.6-fold. Upon DOX introduction, competitive binding between DOX, SG I, and DNA further suppresses background noise, yielding an exceptional fluorescence quenching ratio (F 0 /F) of 52.8. The optimized FAB demonstrates ultra-sensitive DOX detection, with a detection limit of 1.41 nM and a linear response between 1.41 nM and 300 nM. The recoveries of DOX spiked into pond water and drinking water samples ranged from 95.1% to 101.8%, with relative standard deviations (RSDs) below 3.46%, demonstrating negligible matrix effects and confirming the FAB method’s robustness for environmental monitoring applications. This strategy synergistically enhances signal transduction efficiency, and strengthens detection reliability, all while maintaining operational simplicity. As a result, this FAB for DOX detection holds transformative potential, poised to revolutionize biosensor technology in the fields of food safety, environmental monitoring, and biosecurity.