Double Electrochemiluminescence Quenching-Based “Signal-on/off” Immunosensor for CD44 Detection Using Glucose Oxidase-Functionalized CuFe2O4 and Ti3C2Tx@Ni(OH)2-Ru/TPrA Nanoarrays

The rapid and accurate quantification of CD44 in serum is crucial for the early detection and prevention of malignant tumors. In this study, we developed a "signal-on/off" electrochemiluminescence (ECL) immunosensor for the ultrasensitive detection of CD44, leveraging the dual quenching effects of CuFe₂O₄-NH₂-Pd-GOD on Ti₃C₂T_x@Ni(OH)₂-Ru/TPrA nanoarrays. A novel electroactive and catalytic multilayered nanoarray, Ti₃C₂T_x@Ni(OH)₂, was first synthesized and employed as an efficient signal amplifier to accelerate the ECL response by promoting the reaction between Ru(bpy)₃²⁺ and tri-n-propylamine radicals (TPrA^•). To realize effective signal quenching, the electron transfer from Ti₃C₂T_x@Ni(OH)₂-Ru to CuFe₂O₄ occurred. This transfer was driven by appropriate energy level matching and light absorption, leading to significant attenuation of the ECL emission. Moreover, glucose oxidase (GOD)-functionalized CuFe₂O₄ enhanced the in situ generation of H₂O₂, thereby intensifying the quenching effect. The developed immunosensor exhibited a broad linear range from 0.5 pg/mL to 100 ng/mL with an ultralow detection limit of 93 fg/mL. This proposed strategy offers a novel method for the highly sensitive detection of CD44 and holds promise for broader applications in visual biomarker identification.