Electrochemical Biosensor Based on Ultrathin 2D CN-rGO@AgNPs for ROS Detection in Leukemia Cells

Accurate monitoring of ROS levels is essential for oxidative stress (OS) management, but their low concentration and instability pose significant detection challenges. Here, we developed a highly sensitive and selective electrochemical biosensor for real-time, micro-volume ROS detection in leukemia cells. The sensor was fabricated using ultrathin g-C₃N₄-rGO (CN-rGO) nanosheet synthesized through a calcination and hydrothermal approach. These nanosheets were deposited onto a screen-printed carbon electrode (SPE) and further modified with silver nanoparticles (AgNPs) to construct a CN-rGO@AgNPs/SPE biosensor. The porous nanostructure significantly enhanced the catalytic surface area and electron transfer efficiency, achieving high sensitivities towards H₂O₂ and O₂·⁻ detection. Using a window titration platform, the sensor detected ROS in 0.1 mL lysates of leukemia cell lines (U-937, THP-1, and MOLM-13) with concentrations of 7.38 ± 4.81 μM, 11.74 ± 0.985 μM, 19.84 ± 3.73 μM, respectively. Furthermore, the platform allowed precise monitoring of doxorubicin (DOX)-induced OS, providing valuable insights for guiding leukemia chemotherapy. Additionally, Lipopolysaccharide (LPS) stimulation experiments demonstrated the sensor's exceptional real-time performance, enabling dynamic detection of H₂O₂ release kinetics and ROS generation rates at the cellular level. Combining high sensitivity, low detection limits, and micro-volume analysis capabilities, this sensor holds great potential for clinical auxiliary diagnosis and drug efficacy assessment in leukemia research.