Construction of self-enhanced luminescence probes based on Ti₃C₂ reducibility for ultrasensitive PNK analysis

Au nano-clusters (Au NCs) were promising electrochemiluminescence (ECL) nano-materials. However, the small size of Au NCs presented a challenge in terms of their immobilization during the construction of an ECL biosensing platform. This limitation significantly hindered the wider application of Au NCs in the ECL field. In this work, we successfully used the reducibility of Ti₃C₂ to fabricate in situ a self-enhanced nano-probe Ti₃C₂–TiO₂–Au NCs. The strategy of in situ generation not only improved the immobilization of Au NCs on the probe but also eliminated the requirement of adding reducing agents during preparation. In addition, in situ generated TiO₂ could serve as a co-reaction accelerator, shortening the electron transfer distance between S₂O₈²⁻ and Au NCs, thereby improving the utilization of intermediates and enhancing the ECL response of Au NCs. The constructed ECL sensing platform could achieve sensitive detection of polynucleotide kinase (PNK). At the same time, the 5′-end phosphate group of DNA phosphorylation could chelate with a large amount of Ti on the surface of Ti₃C₂, thereby achieving the goal of specific detection of PNK. The sensor based on self-enhanced ECL probes had a broad dynamic range spanning for PNK detection from 10.0 to 1.0 × 10⁷ μU mL⁻¹, with a limit of detection of 1.6 μU mL⁻¹. Moreover, the ECL sensor showed satisfactory detection performance in HeLa cell lysate and serum. This study not only provided insights for addressing the issue of ECL luminescence efficiency in Au NCs but also presented novel concepts for ECL self-enhancement strategies.