Dual-Wavelength Electrochemiluminescence Ratiometric Biosensor for NF-κB p50 Detection with Dimethylthiodiaminoterephthalate Fluorophore and Self-Assembled DNA Tetrahedron Nanostructures Probe

In this work, we fabricated a dual-wavelength electrochemiluminescence ratiometric biosensor based on electrochemiluminescent resonance energy transfer (ECL-RET). In this biosensor, Au nanoparticle-loaded graphitic phase carbon nitride (Au-g-C₃N₄) as a donor and Au-modified dimethylthiodiaminoterephthalate (TAT) analogue (Au@TAT) as an acceptor were investigated for the first time. Besides, tetrahedron DNA probe was immobilized onto Au-g-C₃N₄ to improve the binding efficiency of the transcription factor and ECL ratiometric changes on the basis of the ratio of ECL intensities at 595 and 460 nm, which were obtained through the formation of a sandwich structure of DNA probe–antigen–antibody. Our biosensor achieved the assay of NF-κB p50 with a detection limit of 5.8 pM as well as high stability and specificity.