An electrochemiluminescence-quenched bipolar electrode array for tsRNA assay

<p indent="0mm">Through comparative analysis of high-throughput sequencing data from hepatocellular carcinoma (HCC) pathological tissues and para-carcinoma tissues, we identified differentially expressed genes associated with HCC prognosis, providing a precise and reliable tsRNA target gene, tRF5-29-LysCTT-4, for accurate prognostic evaluation of HCC. Utilizing laser etching and 3D printing technologies, we fabricated a four-channel indium tin oxide (ITO) bipolar electrode (BPE) array. On the cathode surface of the BPE, the target gene tRF5-29-LysCTT-4 initiated a hybridization chain reaction (HCR) to form a DNA polymer that impeded electron transfer, thereby quenching the electrochemiluminescence (ECL) signal of luminol at the BPE anode. This process established a quenching-type multi-channel, multi-sample detection BPE-ECL sensing interface. Under cyclic voltage ranging from 0 to <sc>4 V,</sc> the ECL intensity at the anode exhibited an inverse relationship with the concentration of tRF5-29-LysCTT-4, described by the linear fitting equation <italic>I</italic> = 870.23lg<italic>C</italic>_tsRNA + 8096.38. The detection range spanned from 1.0×10⁻⁵ to 1.0×10² nM, with a detection limit as low as 2.36 fM. In practical assessments of HCC tissue samples, this sensing method was compared with real-time quantitative PCR (RT-qPCR), demonstrating excellent consistency between the two methods. Specifically, the content of tRF5-29-LysCTT-4 was higher in HCC tissues than in the para-carcinoma tissues, indicating the potential clinical application of this BPE-ECL sensing platform for trace detection of tsRNA.