Spherical Nucleic Acid Stabilized Cage Type Three-Dimensional Electrochemiluminescence Probe for Sensitive and...

Accurate nucleic acid-based pathogen diagnosis is critical for clinical treatment and epidemic control. However, present gold-standard testing methods based on nucleic acid amplification are limited by their time and labor-intensive nature. Hence, we reported an amplification-free assay that facilitates rapid, sensitive, and point-of-care detection of infectious diseases. Herein, spherical nucleic acid (SNA)-stabilized cage-type three-dimensional electrochemiluminescence (ECL) reporter probe, constructed utilizing Ru(dcbpy)₃Cl₂-doped NH₂-MIL-101 (Fe) metal-organic framework, was combined with a paper-based bipolar electrode ECL sensor. Furthermore, surface modifications were performed using polydopamine and SNAs to further improve the stability. Notably, compared with the ordinary Ru(dcbpy)₃Cl₂, the modified ECL probe attained 10³-fold enhancement in luminous efficiency. The developed ECL-based nucleic acid biosensing system offers the advantages of simplicity, portability, low cost, and user-friendly nucleic acid detection, with a sample-to-answer time of approximately 15 min. Additionally, the effectiveness of the developed assay in detecting pathogenic nucleic acid was assessed using Mpox and SARS-Cov-2, exhibiting a wide dynamic range from 33 to 10¹⁰ aM and limits of detection as low as 3 copies/µL. Furthermore, the assay exhibited 100% sensitivity and specificity when validated against quantitative polymerase chain reaction-based detection using clinical samples. Altogether, the findings of this study show the efficacy of the novel developed SNA-stabilized cage-type 3D ECL probe-enhanced nucleic acid assay, along with its potential as a promising paradigm for point-of-care diagnosis of infectious pathogens.