Ultrasensitive chemiluminescence assay for the lung cancer biomarker cytokeratin 21-1 via a dual amplification scheme based on the use of encoded gold nanoparticles and a toehold-mediated strand displacement reaction

A catalytic circuit is described in which the addition of a single input DNA strand leads to the release of more than one output strand. The system consists of (a) a three-stranded substrate complex, (b) fuel DNA, and (c) trigger DNA. The whole network is activated by the trigger DNA in the catalytic circuit, which regenerates the trigger to catalyze another new circuit and releases two kinds of bio-barcoded gold nanoparticles (AuNPs) after toehold-mediated strand displacement. The AuNPs are then dissolved to form Au (III) ions which catalyze the luminol-H 2 O 2 chemiluminescence. Cytokeratin 21-1 (CYFRA21-1), associated with so-called non-small cell lung cancer, was used as the model target. By taking advantage of the above dual amplification scheme, this gene assay works in the 20 f. to 8.0 nM CYFRA21-1 concentration range with a 6 f. detection limit. The method exhibits excellent selectivity even over single-mismatched DNA. Schematic of the assay. The addition of target, a lung cancer biomarker, leads to the release of more than one output strand for signal amplification with the detection limit of 6 f. of cytokeratin.