Chemiluminescent detection of nucleic acids induced by peroxidase-like targeted DNA-nanomachines (PxDm) mixed with plasmonic nanoparticles

One of the most critical issues in the field of molecular diagnostics and medicine is the development of compact and sensitive assay devices for the precise detection of nucleic acids. Although there are several effective methods for detecting unique nucleic acid sequences, the high cost of equipment and reagents, as well as the need for highly trained personnel, necessitate the design of new and more affordable diagnostic assays that are comparable in selectivity and sensitivity to existing methods that can be used in developing countries and/or outside of specialized diagnostic laboratories. Sensing methods based on guanine quadruplexes (G-4)/hemin complexes, that have peroxidase activity are one of the promising directions for the detection of target nucleic acids. Target nucleic acid was analyzed by peroxidase-like DNA-nanomachine (PxDm) equipped with 1-3 long analyte binding arms to tightly bind and unwind single-stranded analytes. In this study, we present a technique for sequence-specific detection of nucleic acid. The technique is based on the measuring of a chemiluminescent (CL) emission induced by luminol oxidation utilizing a closed-type detection device. Moreover, the optical properties and potential use of plasmonic silver nanoparticles (Ag NPs) to enhance the CL intensity of chemiluminophore were investigated. Particular attention was paid to the possibility of synthesizing the silver nanoparticles with different spectral positions of plasmon resonance band, depending on the method and duration of synthesis. The CL intensity of luminol in the presence of the post-centrifuged colloidal Ag NPs obtained by laser ablation has been increased 3 times. The combination of AgNPs-luminol-DNA-nanomachine systems in the presence of a target analyte led to the significant increase of limit of detection and reached clinically relevant quantitative indications.