A dual-Color closed bipolar electrochemiluminescence platform for visual simultaneous diagnosis of two pluripotency markers in cancer patients’ urine

Here, for the first time, we introduce a simple yet effective one-step visual approach for the concurrent detection of octamer-binding transcription factor 4 (Oct-4) and Nanog, two pivotal homeobox genes involved in regulating pluripotency in both stem cells and certain cancer cells, within human urine samples. We utilized a closed bipolar electrochemistry system to enable the electrochemiluminescence (ECL) detection of pluripotency biomarkers. This was accomplished by capturing the ECL of Ru(bpy)₃²⁺ loaded into mesoporous SiO₂ nanoparticles and luminol incorporated into luminol@MIL-53 on the upper and lower anodic poles of BPEs in separate chambers of a bipolar cell using a digital camera. An identical electric potential was applied across the bipolar electrodes, driving the reduction of thionine acetate at the cathodic poles and thereby facilitating simultaneous light emission of the luminophores. Moreover, the incorporation of electroactive Ti₃C₂T_x MXene-TiO₂ nanosheets catalyzed the electro-oxidation of co-reactants within both the Ru(bpy)₃²⁺/TPrA and luminol/H₂O₂ systems, thereby enhancing the signal-to-background ratio and eliciting a significantly amplified visual ECL response. Under optimized conditions, the immunosensor displayed a linear response across a broad range of 100 pg mL⁻¹ to 400 ng/mL for Nanog, with a detection limit of 21.58 pg mL⁻¹, and a linear response from 200 pg mL⁻¹ to 400 ng/mL with a detection limit of 106.24 pg mL⁻¹ for Oct-4. This study supports the immunosensor's potential for broader clinical application, where its high specificity and sensitivity could offer significant benefits in cancer diagnostics and stem cell research.